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The Hedgehog (Hh) signaling pathway is involved in T cell differentiation and development and plays a major regulatory part in different stages of T cell development. A previous study by us suggested that prenatal exposure to staphylococcal enterotoxin B (SEB) changed the percentages of T cell subpopulation in the offspring thymus. However, it is unclear whether prenatal SEB exposure impacts the Hh signaling pathway in thymic T cells. In the present study, pregnant rats at gestational day 16 were intravenously injected once with 15 µg SEB, and the thymi of both neonatal and adult offspring rats were aseptically acquired to scrutinize the effects of SEB on the Hh signaling pathway. It firstly found that prenatal SEB exposure clearly caused the increased expression of Shh and Dhh ligands of the Hh signaling pathway in thymus tissue of both neonatal and adult offspring rats, but significantly decreased the expression levels of membrane receptors of Ptch1 and Smo, transcription factor Gli1, as well as target genes of CyclinD1, C-myc, and N-myc in Hh signaling pathway of thymic T cells. These data suggest that prenatal SEB exposure inhibits the Hh signaling pathway in thymic T lymphocytes of the neonatal offspring, and this effect can be maintained in adult offspring via the imprinting effect.
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Enterotoxinas , Proteínas Hedgehog , Transducción de Señal , Linfocitos T , Timo , Animales , Proteínas Hedgehog/metabolismo , Proteínas Hedgehog/genética , Femenino , Embarazo , Ratas , Timo/metabolismo , Timo/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Proteína con Dedos de Zinc GLI1/metabolismo , Proteína con Dedos de Zinc GLI1/genética , Receptor Patched-1/metabolismo , Receptor Patched-1/genética , Receptor Smoothened/metabolismo , Receptor Smoothened/genética , Efectos Tardíos de la Exposición Prenatal/inmunología , Diferenciación Celular/efectos de los fármacos , Ratas Sprague-Dawley , MasculinoRESUMEN
Hedgehog (Hh) signaling plays a significant role in embryogenesis and several physiological processes, such as wound healing and organ homeostasis. In a pathological setting, it is associated with oncogenesis and is responsible for disease progression and poor clinical outcomes. Hedgehog signaling mediates downstream actions via Glioma Associated Oncogene Homolog (GLI) transcription factors. Inhibiting Hh signaling is an important oncological strategy in which inhibitors of the ligands SMO or GLI have been looked at. This review briefly narrates the Hh ligands, signal transduction, the target genes involved and comprehensively describes the numerous inhibitors that have been evaluated for use in various neoplastic settings.
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Proteínas Hedgehog , Neoplasias , Humanos , Transducción de Señal/genética , Alcaloides de Veratrum , Neoplasias/tratamiento farmacológicoRESUMEN
Lenvatinib is a frontline tyrosine kinase inhibitor for patients with advanced hepatocellular carcinoma (HCC). However, just 25% of patients benefit from the treatment, and acquired resistance always develops. To date, there are neither effective medications to combat lenvatinib resistance nor accurate markers that might predict how well a patient would respond to the lenvatinib treatment. Thus, novel strategies to recognize and deal with lenvatinib resistance are desperately needed. In the current study, a robust Lenvatinib Resistance index (LRi) model to predict lenvatinib response status in HCC was first established. Subsequently, five candidate drugs (Mercaptopurine, AACOCF3, NU1025, Fasudil, and Exisulind) that were capable of reversing lenvatinib resistance signature were initially selected by performing the connectivity map (CMap) analysis, and fasudil finally stood out by conducting a series of cellular functional assays in vitro and xenograft mouse model. Transcriptomics revealed that the co-administration of lenvatinib and fasudil overcame lenvatinib resistance by remodeling the hedgehog signaling pathway. Mechanistically, the feedback activation of EGFR by lenvatinib led to the activation of the GLI2-ABCC1 pathway, which supported the HCC cell's survival and proliferation. Notably, co-administration of lenvatinib and fasudil significantly inhibited IHH, the upstream switch of the hedgehog pathway, to counteract GLI2 activation and finally enhance the effectiveness of lenvatinib. These findings elucidated a novel EGFR-mediated mechanism of lenvatinib resistance and provided a practical approach to overcoming drug resistance in HCC through meaningful drug repurposing strategies.
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1-(5-Isoquinolinesulfonil)-2-Metilpiperazina/análogos & derivados , Antineoplásicos , Carcinoma Hepatocelular , Neoplasias Hepáticas , Compuestos de Fenilurea , Quinolinas , Humanos , Animales , Ratones , Carcinoma Hepatocelular/metabolismo , Proteínas Hedgehog , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/metabolismo , Línea Celular Tumoral , Receptores ErbB , Proteína Gli2 con Dedos de Zinc , Proteínas NuclearesRESUMEN
The Hedgehog (Hh) signaling pathway plays important roles in various physiological functions. Several malignancies, such as basal cell carcinoma (BCC) and medulloblastoma (MB), have been linked to the aberrant activation of Hh signaling. Although therapeutic drugs have been developed to inhibit Hh pathway-dependent cancer growth, drug resistance remains a major obstacle in cancer treatment. Here, we show that the newly identified, 2-{3-[1-(benzylsulfonyl)-1,2,3,6-tetrahydropyridin-4-yl]-2-methyl-1H-indol-1-yl}-1-(pyrrolidin-1-yl)ethenone analog (LKD1214) exhibits comparable potency to vismodegib in suppressing the Hh pathway activation. LKD1214 represses Smoothened (SMO) activity by blocking its ciliary translocation. Interestingly, we also identified that it has a distinctive binding interface with SMO compared with other SMO-regulating chemicals. Notably, it maintains an inhibitory activity against the SmoD477H mutant, as observed in a patient with vismodegib-resistant BCC. Furthermore, LKD1214 inhibits tumor growth in the mouse model of MB. Collectively, these findings suggest that LKD1214 has the therapeutic potential to overcome drug-resistance in Hh-dependent cancers.
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Hedgehog (Hh) signaling is crucial in cardiovascular development and maintenance. However, the biological role of Patched1 (Ptch1), an inhibitory receptor of the Hh signaling pathway, remains elusive. In this study, a Ptch1 ortholog was characterized in Nile tilapia (Oreochromis niloticus), and its function was investigated through CRISPR/Cas9 gene knockout. When one-cell embryos were injected with CRISPR/Cas9 targeting ptch1, the mutation efficiency exceeded 70%. During 0-3 days post fertilization (dpf), no significant differences were observed between the ptch1 mutant group and the control group; at 4 dpf (0 day after hatching), about 10% of the larvae showed an angiogenesis defect and absence of blood flow; from 5 dpf, most larvae exhibited an elongated heart, large pericardial cavity, and blood leakage and coagulation, ultimately dying during the 6-8 dpf period due to the lack of blood circulation. Consistently, multiple differentially expressed genes related to angiogenesis, blood coagulation, and heart development were enriched in the ptch1 mutants. Furthermore, Smoothened (Smo) antagonist (cyclopamine) treatment of the ptch1 mutants greatly rescued the cardiovascular disorders. Collectively, our study suggests that Ptch1 is required for cardiovascular development and vascular integrity via Smo signaling, and excessive Hh signaling is detrimental to cardiovascular development.
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Cíclidos , Animales , Cíclidos/metabolismo , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Transducción de Señal , Técnicas de Inactivación de Genes , Mutación , Receptor Smoothened/genéticaRESUMEN
INTRODUCTION: Parkinson's disease (PD) is the most common neurodegenerative disease worldwide. Studies have shown that insulin-like growth factor-binding protein 5 (IGFBP5) may contribute to methamphetamine-induced neurotoxicity and neuronal apoptosis in PC-12 cells and rat striatum. Here, we studied the expression and role of IGFBP5 in the 6-OHDA-toxicant model of PD. METHODS: PC-12 and SH-SY5Y cells were exposed to 50 µ
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Apoptosis , Proteínas Hedgehog , Proteína 5 de Unión a Factor de Crecimiento Similar a la Insulina , Oxidopamina , Enfermedad de Parkinson , Transducción de Señal , Animales , Proteínas Hedgehog/metabolismo , Ratas , Apoptosis/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Humanos , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Modelos Animales de Enfermedad , Masculino , Células PC12 , Ratas Sprague-DawleyRESUMEN
Changes in protein ubiquitination have been linked to cancer. Deubiquitinating enzymes (DUBs) counteract E3 ligase activities and have emerged as promising targets for cancer treatment. Ubiquitin-specific peptidase 5 (USP5) is a member of the DUBs family and has been implicated in promoting tumorigenesis in numerous cancers. However, the clinical significance and biological function of USP5 in osteosarcoma (OS) remains unclear. Here, we found elevated USP5 expression in OS tissues compared with normal bone tissues. Furthermore, we observed significant associations of elevated USP5 levels with increased mortality and more malignant phenotypes in OS patients. Moreover, our results revealed that USP5 could facilitate metastasis and cell progression in OS by activating the hedgehog (Hh) signaling pathway using cultured cells and animal tumor models. Mechanistically, USP5 appeared to stabilize and deubiquitinate Gli1, a key mediator of the Hh signaling pathway. Additionally, the oncogenic effect of USP5 in OS was dependent on Gli1 stability. Our findings support the model where USP5 contributes to OS pathogenesis by activating the Hh/Gli1 signaling pathway, making USP5 a potential diagnostic and therapeutic target for OS.
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Approximately a quarter of Retinitis Pigmentosa (RP) is caused by mutations in transport-related genes in cilia. IFT27 (Intraflagellar Transport 27), a core component of the ciliary intraflagellar transport (IFT) system, has been implicated as a significant pathogenic gene in RP. The pathogenic mechanisms and subsequent pathology related to IFT27 mutations in RP are largely obscure. Here, we utilized TALEN technology to create an ift27 knockout (ift27-/-) zebrafish model. Electroretinography (ERG) detection showed impaired vision in this model. Histopathological examinations disclosed that ift27 mutations cause progressive degeneration of photoreceptors in zebrafish, and this degeneration was late-onset. Immunofluorescence labeling of outer segments showed that rods degenerated before cones, aligning with the conventional characterization of RP. In cultured human retinal pigment epithelial cells, we found that IFT27 was involved in maintaining ciliary morphology. Furthermore, decreased IFT27 expression resulted in the inhibition of the Hedgehog (Hh) signaling pathway, including decreased expression of key factors in the Hh pathway and abnormal localization of the ciliary mediator Gli2. In summary, we generated an ift27-/- zebrafish line with retinal degeneration which mimicked the symptoms of RP patients, highlighting IFT27's integral role in the long-term maintenance of cilia via the Hh signaling pathway. This work may furnish new insights into the treatment or delay of RP caused by IFT27 mutations.
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Retinitis Pigmentosa , Proteínas de Pez Cebra , Pez Cebra , Animales , Humanos , Transporte Biológico , Cilios/genética , Proteínas Hedgehog/genética , Proteínas Hedgehog/metabolismo , Retinitis Pigmentosa/genética , Retinitis Pigmentosa/patología , Pez Cebra/genética , Pez Cebra/metabolismo , Proteínas de Pez Cebra/genéticaRESUMEN
Multipotent mesenchymal stem cells (MSCs) have high self-renewal and multi-lineage differentiation potentials and low immunogenicity, so they have attracted much attention in the field of regenerative medicine and have a promising clinical application. MSCs originate from the mesoderm and can differentiate not only into osteoblasts, cartilage, adipocytes, and muscle cells but also into ectodermal and endodermal cell lineages across embryonic layers. To design cell therapy for replacement of damaged tissues, it is essential to understand the signaling pathways, which have a major impact on MSC differentiation, as this will help to integrate the signaling inputs to initiate a specific lineage. Hedgehog (Hh) signaling plays a vital role in the development of various tissues and organs in the embryo. As a morphogen, Hh not only regulates the survival and proliferation of tissue progenitor and stem populations but also is a critical moderator of MSC differentiation, involving tri-lineage and across embryonic layer differentiation of MSCs. This review summarizes the role of Hh signaling pathway in the differentiation of MSCs to mesodermal, endodermal, and ectodermal cells.
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Diferenciación Celular , Proteínas Hedgehog , Células Madre Mesenquimatosas , Transducción de Señal , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/citología , Proteínas Hedgehog/metabolismo , Humanos , Diferenciación Celular/fisiología , Animales , Células Madre Multipotentes/citología , Células Madre Multipotentes/metabolismoRESUMEN
The Hedgehog (Hh) signaling pathway plays a crucial role in diverse biological processes such as cell differentiation, proliferation, senescence, tumorigenesis, malignant transformation, and drug resistance. Aberrant Hh signaling, resulting from mutations and excessive activation, can contribute to the development of various diseases during different stages of biogenesis and development. Moreover, it has been linked to unfavorable outcomes in several human cancers, including basal cell carcinoma (BCC), multiple myeloma (MM), melanoma, and breast cancer. Hence, the presence of mutations and excessive activation of the Hh pathway presents obstacles and constraints in the realm of cancer treatment. Extant research has demonstrated that small molecule inhibitors are regarded as the most effective therapeutic approaches for targeting the Hh pathway in contrast to traditional chemotherapy and radiotherapy. Consequently, this review focuses on the present repertoire of small molecule inhibitors that target various components of the Hh pathway, including Hh ligands, Ptch receptors, Smo transmembrane proteins, and Gli nuclear transcription factors. This study provides a comprehensive analysis of small molecules' structural and functional aspects in the preclinical and clinical management of cancer. Additionally, it elucidates the obstacles encountered in targeting the Hh pathway for human cancer therapy and proposes potential therapeutic approaches.
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Antineoplásicos , Proteínas Hedgehog , Neoplasias , Transducción de Señal , Humanos , Proteínas Hedgehog/antagonistas & inhibidores , Proteínas Hedgehog/metabolismo , Transducción de Señal/efectos de los fármacos , Antineoplásicos/farmacología , Antineoplásicos/química , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Neoplasias/patología , Bibliotecas de Moléculas Pequeñas/farmacología , Bibliotecas de Moléculas Pequeñas/química , AnimalesRESUMEN
HCC remains one of the leading causes of cancer-related death globally. The main challenges in treatments of hepatocellular carcinoma (HCC) primarily arise from high rates of postoperative recurrence and the limited efficacy in treating advanced-stage patients. Various signaling pathways involved in HCC have been reported. Among them, the Sonic hedgehog (SHH) signaling pathway is crucial. The presence of SHH ligands is identified in approximately 60% of HCC tumor tissues, including tumor nests. PTCH-1 and GLI-1 are detected in more than half of HCC tissues, while GLI-2 is found in over 84% of HCC tissues. The SHH signaling pathway (including canonical and non-canonical) is involved in different aspects of HCC, including hepatocarcinogenesis, tumor growth, tumor invasiveness, progression, and migration. The SHH signaling pathway also contributes to recurrence, metastasis, modulation of the cancer microenvironment, and sustaining cancer stem cells. It also affects the resistance of HCC cells to chemotherapy, target therapy, and radiotherapy. Reappraisal of the roles of the SHH signaling pathway in HCC may trigger some novel therapies for HCC.
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OBJECTIVE: Rheumatoid arthritis (RA) is a chronic, progressive autoimmune disease with a complex pathogenesis that has not yet been fully elucidated, and T-cell pyroptosis is an important pathogenetic factor in RA. This study aimed to investigate the role of endoplasmic reticulum aminopeptidase 2 (ERAP2) in the pyroptosis of CD4+ T cells in RA and the specific molecular mechanism. METHODS: Peripheral venous blood was collected from human subjects, and CD4+ T cells were isolated and activated to measure the level of pyroptosis and ERAP2 expression. Pyroptosis levels were assessed using immunofluorescence, flow cytometry, qRT-PCR, and Western blotting. Changes in pyroptosis levels were observed upon knockdown or overexpression of ERAP2. To detect activated Caspase-1 in tissues, chimeric mice were engrafted with human synovial tissue and reconstituted with human CD4+ T cells. CD4 + T cells were treated with GLI1 antagonists and SMO receptor agonists to detect changes in pyroptosis levels. RESULTS: CD4+ T cell levels undergoing pyroptosis were found to be elevated in the blood and synovium of RA patients. The gene and protein expression of ERAP2 were significantly higher in CD4+ T cells from RA patients. Deletion of ERAP2 suppressed pyroptosis of these cells, attenuated the activation of Caspase-1 in tissue T cells, and reduced tissue inflammatory responses. Reciprocally, overexpression of ERAP2 triggered inflammasome assembly, activated Caspase-1, and induced pyroptosis in CD4+ T cells. Mechanistically, ERAP2 inhibits the Hedgehog signaling pathway and upregulates the expression of nucleotide-binding oligomerization segment-like receptor family 3(NLRP3), cleaved Caspase-1, and Gasdermin D to promote pyroptosis in CD4+ T cells. CONCLUSIONS: Taken together, our results identify a novel mechanism by which ERAP2 regulates RA development and document the effect of the ERAP2/Hedgehog signaling axis on pyroptosis of CD4+ T cells from RA patients.
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Artritis Reumatoide , Piroptosis , Humanos , Animales , Ratones , Proteínas Hedgehog/metabolismo , Artritis Reumatoide/metabolismo , Linfocitos T CD4-Positivos , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Inflamasomas/metabolismo , Caspasa 1/metabolismo , Aminopeptidasas/genética , Retículo Endoplásmico/metabolismo , Retículo Endoplásmico/patologíaRESUMEN
Veratrum californicum contains steroidal alkaloids that function as inhibitors of hedgehog (Hh) signaling, a pathway involved in the growth and differentiation of cells and normal tissue development. This same Hh pathway is abnormally active for cell proliferation in more than 20 types of cancer. In this current study, alkaloids have been extracted from the root and rhizome of V. californicum, followed by their separation into five fractions using high performance liquid chromatography. Mass spectrometry was used to identify the presence of twenty-five alkaloids, nine more than are commonly cited in literature reports, and the Bruker Compass Data Analysis software was used to predict the molecular formula for every detected alkaloid. The Gli activity of the raw extract and each fraction were compared to 0.1 µM cyclopamine, and fractions 1, 2, and 4 showed increased bioactivity through suppression of the Hh signaling pathway. Fractions 2 and 4 had enhanced bioactivity, but fraction 1 was most effective in inhibiting Hh signaling. The composition of fraction 1 consisted of veratrosine, cycloposine, and potential isomers of each.
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Orofacial clefts (OFCs) represent the most prevalent congenital craniofacial anomalies, significantly impacting patients' appearance, oral function, and psychological well-being. Among these, non-syndromic OFCs (NSOFCs) are the most predominant type, with the etiology attributed to a combination of genetic and environmental factors. Rare variants of key genes involved in craniofacial development-related signaling pathway are crucial in the occurrence of NSOFCs, and our recent studies have identified PTCH1, a receptor-coding gene in the Hedgehog signaling pathway, as a causative gene for NSOFCs. However, the role of PTCH2, the paralog of PTCH1, in pathogenesis of NSOFCs remains unclear. Here, we perform whole-exome sequencing to explore the genetic basis of 144 sporadic NSOFC patients. We identify five heterozygous variants of PTCH2 in four patients: p.L104P, p.A131G, p.R557H, p.I927S, and p.V978D, with the latter two co-occurring in a single patient. These variants, all proven to be rare through multiple genomic databases, with p.I927S and p.V978D being novel variants and previously unreported. Sequence alignment suggests that these affected amino acids are evolutionarily conserved across vertebrates. Utilizing predictive structural modeling tools such as AlphaFold and SWISS-MODEL, we propose that these variants may disrupt the protein's structure and function. In summary, our findings suggest that PTCH2 may be a novel candidate gene predicted to be associated with NSOFCs, thereby broadening the spectrum of causative genes implicated in the craniofacial anomalies.
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Labio Leporino , Fisura del Paladar , Receptor Patched-2 , Animales , Humanos , Encéfalo/anomalías , Labio Leporino/genética , Fisura del Paladar/genética , Proteínas Hedgehog/genética , Receptor Patched-2/genética , Transducción de SeñalRESUMEN
Oral squamous cell carcinoma (OSCC) represents the primary subtype of head and neck squamous cell carcinoma (HNSCC), characterized by a high morbidity and mortality rate. Although previous studies have established specific correlations between euchromatic histone lysine methyltransferase 2 (EHMT2), a histone lysine methyltransferase, and the malignant phenotype of OSCC cells, its biological functions in OSCC remain largely unknown. This study, grounded in bioinformatics predictions, aims to clarify the influence of EHMT2 on the malignant behavior of OSCC cells and delve into the underlying mechanisms. EHMT2 exhibited high expression in OSCC tissues and demonstrated an association with poor patient outcomes. Artificial EHMT2 silencing in OSCC cells, achieved through lentiviral vector infection, significantly inhibited colony formation, migration, invasion, and cell survival. Regarding the mechanism, EHMT2 was found to bind the promoter of arrestin beta 1 (ARRB1), thereby suppressing its transcription through H3K9me2 modification. ARRB1, in turn, was identified as a negative regulator of the Hedgehog pathway, leading to a reduction in the proteins GLI1 and PTCH1. Cancer stem cells (CSCs) were enriched through repeated sphere formation assays in two OSCC cell lines. EHMT2 was found to activate the Hedgehog pathway, thus promoting sphere formation, migration and invasion, survival, and tumorigenic activity of the OSCC-CSCs. Notably, these effects were counteracted by the additional overexpression of ARRB1. In conclusion, this study provides novel evidence suggesting that EHMT2 plays specific roles in enhancing stem cell properties in OSCC by modulating the ARRB1-Hedgehog signaling cascade.
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Asthma is the most prevalent chronic inflammatory disease of the airways in children. The most prevalent phenotype of asthma is eosinophilic asthma, which is driven by a Th2 immune response and can be effectively managed by inhaled corticosteroid therapy. However, there are phenotypes of asthma with Th17 immune response that are insensitive to corticosteroid therapy and manifest a more severe phenotype. The treatment of this corticosteroid-insensitive asthma is currently immature and requires further attention. The objective of this study is to elucidate the regulation of the Hedgehog signaling pathway in Th17 cell differentiation in asthma. The study demonstrated that both Smo and Gli3, key components of the Hedgehog signaling pathway, were upregulated in Th17 polarization in vitro and in a Th17-dominant asthma model in vivo. Inhibiting Smo with a small molecule inhibitor or genetically knocking down Gli3 was found to suppress Th17 polarization. Smo was found to increase in Th1, Th2, Th17 and Treg polarization, while Gli3 specifically increased in Th17 polarization. ChIP-qPCR analyses indicated that Gli3 can directly interact with IL-6 in T cells, inducing STAT3 phosphorylation and promoting Th17 cell differentiation. Furthermore, the study demonstrated a correlation between elevated Gli3 expression and IL-17A and IL-6 expression in children with asthma. In conclusion, the study demonstrated that the Hedgehog signaling pathway plays an important role in the pathogenesis of asthma, as it regulates the differentiation of Th17 cells through the IL-6/STAT3 signaling. This may provide a potential therapeutic target for corticosteroid-insensitive asthma driven by Th17 cells.
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Asma , Diferenciación Celular , Proteínas Hedgehog , Interleucina-6 , Factor de Transcripción STAT3 , Transducción de Señal , Células Th17 , Proteína Gli3 con Dedos de Zinc , Asma/inmunología , Asma/metabolismo , Asma/tratamiento farmacológico , Células Th17/inmunología , Factor de Transcripción STAT3/metabolismo , Animales , Interleucina-6/metabolismo , Diferenciación Celular/efectos de los fármacos , Proteínas Hedgehog/metabolismo , Humanos , Proteína Gli3 con Dedos de Zinc/metabolismo , Proteína Gli3 con Dedos de Zinc/genética , Ratones , Niño , Masculino , Receptor Smoothened/metabolismo , Receptor Smoothened/genética , Femenino , Ratones Endogámicos BALB C , Modelos Animales de Enfermedad , Células Cultivadas , Proteínas del Tejido NerviosoRESUMEN
Background: The liver's regenerative capacity allows it to repair itself after injury. Extracellular vesicles and particles (EVPs) in the liver's interstitial space are crucial for signal transduction, metabolism, and immune regulation. Understanding the role and mechanism of liver-derived EVPs in regeneration is significant, particularly after partial hepatectomy, where the mechanisms remain unclear. Methods: A 70% hepatectomy model was established in mice, and EVPs were isolated and characterized using electron microscopy, nanocharacterization, and Western blot analysis. Combined metabolomic and transcriptomic analyses revealed ß-sitosterol enrichment in EVPs and activation of the Hedgehog signaling pathway during regeneration. The role of ß-sitosterol in EVPs on the Hedgehog pathway and its targets were identified using qRT-PCR, Western blot analysis. The regulation of carnitine synthesis by this pathway was determined using a dual luciferase assay. The effect of a ß-sitosterol diet on liver regeneration was verified in mice. Results: After 70% hepatectomy, the liver successfully regenerated without liver failure or death. At 24 hours post-surgery, tissue staining showed transient regeneration-associated steatosis (TRAS), with increased Ki67 positivity at 48 hours. EVPs displayed a spherical lipid bilayer structure with particle sizes of 70-130 nm. CD9, CD63, and CD81 in liver-derived EVPs were confirmed. Transcriptomic and metabolomic analyses showed EVPs supplementation significantly promoted carnitine synthesis and fatty acid oxidation. Tissue staining confirmed accelerated TRAS resolution and enhanced liver regeneration with EVP supplementation. Mass spectrometry identified ß-sitosterol in EVPs, which binds to Smo protein, activating the Hedgehog pathway. This led to the nuclear transport of Gli3, stimulating Setd5 transcription and inducing carnitine synthesis, thereby accelerating fatty acid oxidation. Mice with increased ß-sitosterol intake showed faster TRAS resolution and liver regeneration compared to controls. Conclusion: Liver-derived EVPs promote regeneration after partial hepatectomy. ß-sitosterol from EVPs accelerates fatty acid oxidation and promotes liver regeneration by activating Hedgehog signaling pathway.
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Vesículas Extracelulares , Proteínas Hedgehog , Hepatectomía , Regeneración Hepática , Hígado , Sitoesteroles , Animales , Sitoesteroles/farmacología , Sitoesteroles/química , Regeneración Hepática/efectos de los fármacos , Regeneración Hepática/fisiología , Vesículas Extracelulares/efectos de los fármacos , Vesículas Extracelulares/química , Ratones , Hígado/efectos de los fármacos , Hígado/metabolismo , Proteínas Hedgehog/metabolismo , Masculino , Transducción de Señal/efectos de los fármacos , Ratones Endogámicos C57BL , Carnitina/farmacología , Tamaño de la PartículaRESUMEN
Hedgehog (Hh) signaling pathway dysregulation is involved in the pathogenesis of metabolic dysfunction-associated steatohepatitis, and the sonic Hh (SHh) protein, a pivotal molecule in the Hh pathway, is expressed in ballooned hepatocytes. The present study aimed to investigate the clinicopathological significance of SHh expression in steatohepatitic hepatocellular carcinoma (SH-HCC). Reverse transcription-quantitative polymerase chain reaction and immunohistochemistry were performed to examine SHh gene and SHh protein expression in SH-HCC. Additionally, patients with conventional HCC (C-HCC) were included in the control group. Comparisons of patient and tumor characteristics were also performed. The prevalence of SH-HCC was 3% in the whole cohort, and it was significantly associated with a high prevalence of diabetes mellitus. SHh mRNA was detected in all patients with SH-HCC, but not in 23% of patients with C-HCC. Notably, SHh mRNA expression was not significantly different between patients with SH-HCC and those with C-HCC; however, high SHh protein expression was significantly more frequent in SH-HCC patients than in those with C-HCC. Although the prognosis was not significantly different between the SH-HCC and C-HCC groups, high SHh protein expression was an independent poor prognostic factor for HCC. In conclusion, SHh could potentially serve as a therapeutic target for patients with HCC.
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The Hedgehog (Hh) signaling pathway plays an important role in the development and progression of hepatocellular carcinoma and its tumor microenvironment, and abnormal activation of Hh signal can accelerate the growth of tumor. The crosstalk between the Hh signaling pathway and TME is closely associated with tumor growth and the formation of inhibitory tumor microenvironment. Evidence shows that inhibition of Hh signal plays an important role in inhibiting the growth of hepatocellular carcinoma. This article reviews the current research status of the role, mechanism, and potential therapeutic significance of abnormal activation of Hh signal in hepatocellular carcinoma and its tumor microenvironment, so as to provide new ideas for the treatment of hepatocellular carcinoma.
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Objective To compare the pathological status of gastric mucosa and the expression of HH-PTCH-SMO-GLI(Hedgehog signaling pathway)and NOX/NF-κB/STAT1 signaling pathways in Hp and non-HP infected CAG patients,and to explore the biological mechanism of Hp promoting the"inflammatory cancer transformation"of CAG.Methods 43 patients with CAG who met the criteria were enrolled and divided into CAG with Hp infection group(Hp+ CAG group,n=21)and CAG without Hp infection group(HP-CAG group,n=22).The histological changes of gastric mucosa were observed by hematoxylin-eosin(HE)staining.Western blot was used to detect the relative expression levels of NOX1,NOX2,NOX4,STAT1,P65 and P-P65 in gastric mucosa.Real-time fluorescence quantitative PCR(RT-qPCR)was used to detect Gli1 mRNA,Gli2 mRNA,Gli3 mRNA,Shh mRNA,Smo mRNA,Ptch mRNA,NOX1 mRNA,NOX2 mRNA,NOX4 mRNA and NF-κB mRNA in gastric mucosa The mRNA level.Results HE staining results of gastric tissues in the two groups:In the Hp+CAG group,gastric epithelial cells were partially necrotic and shed,the surface was not smooth,the number of glands was reduced and disordered,intestinal metaplasia was observed,and diffuse lymphocyte and neutrophil infiltration were observed in the lamina proper.The degree of lymphocyte and neutrophil infiltration in HP-CAG group was lighter than that in Hp+CAG group.RT-qPCR results:Compared with HP-CAG group,the levels of Gli1 mRNA,Shh mRNA,Smo mRNA and Ptch mRNA in gastric mucosa of Hp+CAG group were significantly decreased(P<0.01).The levels of Gli2 mRNA,Gli3 mRNA,NOX1 mRNA,NOX2 mRNA,NOX4 mRNA and NF-κB mRNA were significantly increased(P<0.01).Western blot detection results:Compared with hP-CAG group,the relative expression levels of NOX1/GAPDH,NOX2/GAPDH,NOX4/GAPDH and P-P65/GAPDH in gastric mucosa of Hp+CAG group were significantly increased(P<0.01),and the STAT1 level was significantly decreased(P<0.01).There was no significant difference in the relative expression of P65/GAPDH between the two groups(P>0.05).Conclusion Hp infection may cause long-term inflammation of gastric mucosa,promote atrophy and intestinal metaplasia,and increase the risk of cancer by inhibiting hH-PTC-SMO-GLi signaling pathway and abnormal activation of NOX/NF-κB/STAT1 signaling pathway.