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Objective: We aimed to investigate the hypothesis that interferon (IFN)–stimulated gene (ISG) expression in systemic lupus erythematosus (SLE) monocytes is linked to changes in metabolic reprogramming and epigenetic regulation of ISG expression. Methods: Monocytes from healthy volunteers and patients with SLE at baseline or following IFNα treatment were analyzed by extracellular flux analysis, proteomics, metabolomics, chromatin immunoprecipitation, and gene expression. The histone demethylases KDM6A/B were inhibited using glycogen synthase kinase J4 (GSK‐J4). GSK‐J4 was tested in pristane and resiquimod (R848) models of IFN‐driven SLE. Results: SLE monocytes had enhanced rates of glycolysis and oxidative phosphorylation compared to healthy control monocytes, as well as increased levels of isocitrate dehydrogenase and its product, α‐ketoglutarate (α‐KG). Because α‐KG is a required cofactor for histone demethylases KDM6A and KDM6B, we hypothesized that IFNα may be driving "trained immune" responses through altering histone methylation. IFNα priming (day 1) resulted in a sustained increase in the expression of ISGs in primed cells (day 5) and enhanced expression on restimulation with IFNα. Importantly, decreased H3K27 trimethylation was observed at the promoters of ISGs following IFNα priming. Finally, GSK‐J4 (KDM6A/B inhibitor) resulted in decreased ISG expression in SLE patient monocytes, as well as reduced autoantibody production, ISG expression, and kidney pathology in R848‐treated BALB/c mice. Conclusion: Our study suggests long‐term IFNα exposure alters the epigenetic regulation of ISG expression in SLE monocytes via changes in immunometabolism, a mechanism reflecting trained immunity to type I IFN. Importantly, it opens the possibility that targeting histone‐modifying enzymes, such as KDM6A/B, may reduce IFN responses in SLE. [ABSTRACT FROM AUTHOR]

Objective Sialylation of the crystallizable fragment (Fc) of ACPAs, which is catalysed by β-galactoside α-2,6-sialyltransferase 1 (ST6GAL1) could attenuate inflammation of RA. In this study, we screened the transcription factor of ST6GAL1 and elucidated the mechanism of transcriptionally upregulating sialylation of ACPAs in B cells to explore its role in the progression of RA. Methods Transcription factors interacting with the P2 promoter of ST6GAL1 were screened by DNA pull-down and liquid chromatography with tandem mass spectrometry (LC-MS/MS), and verified by chromatin immunoprecipitation (ChIP), dual luciferase reporter assay and electrophoretic mobility shift assay (EMSA). The function of the CCCTC-binding factor (CTCF) on the expression of ST6GAL1 and the inflammatory effect of ACPAs were verified by knocking down and overexpressing CTCF in B cells. The CIA model was constructed from B cell–specific CTCF knockout mice to explore the effect of CTCF on arthritis progression. Results We observed that the levels of ST6GAL1 and ACPAs sialylation decreased in the serum of RA patients and were negatively correlated with DAS28 scores. Subsequently, CTCF was screened and verified as the transcription factor interacting with the P2 promoter of ST6GAL1, which enhances the sialylation of ACPAs, thus weakening the inflammatory activity of ACPAs. Furthermore, the above results were also verified in the CIA model constructed from B cell–specific CTCF knockout mice. Conclusion CCCTC-binding factor is the specific transcription factor of β-galactoside α-2,6-sialyltransferase 1 in B cells that upregulates the sialylation of ACPAs in RA and attenuates the disease progression. [ABSTRACT FROM AUTHOR]

Objectives: In vivo, the principal function of mechanosensitive odontoblasts is to synthesize and secrete the matrix which then calcifies and forms reactive dentin after exposure to appropriate stimuli. This study aims to develop the influence of mechanical factors on dentinogenesis based on odontoblasts, which contribute to reparative dentin formation. Methods: We fabricated polydimethylsiloxane with different stiffnesses and seeded 17IIA11 odontoblast‐like cells on the substrates in different stiffnesses. Cell morphology was detected by scanning electron microscope, and the mineralization phenotype was detected by alkaline phosphatase staining and alizarin red staining, while expression levels of dentinogenesis‐related genes (including Runx2, Osx, and Alp) were assayed by qPCR. To explore mechanism, protein distribution and expression levels were detected by immunofluorescent staining, Western blotting, and immunoprecipitation. Results: In our results, during dentinogenesis, 17IIA11 odontoblast‐like cells appeared better extension on stiffer substrates. The binding between LAMB1 and FAK contributed to converting mechanical stimuli into biochemical signaling, thereby controlling mitogen‐activated protein kinase kinase 1/2 activity in stiffness‐driven dentinogenesis. Conclusion: The present study suggests odontoblast behaviors can be directly regulated by mechanical factors at cell–material interfaces, which offers fundamental mechanism in remodeling cell microenvironment, thereby contributing to physiological phenomena explanation and tissue engineering progress. [ABSTRACT FROM AUTHOR]

Objective. This study was to construct a ceRNA (lncRNA-miRNA-mRNA) regulatory network for prostate cancer (PCa) and to explore the prognostic correlation, key biological functions, and pathways of core RNAs. Methods. Three subgene expression matrices (miRNA, lncRNA, and mRNA expression matrix) were taken from the TCGA database and used in this investigation. Differential expression analysis and differential expression miRNAs were carried out. Next, the ceRNA (lncRNA-miRNA-mRNA) regulatory complex was used to visualize our data and show how they interacted. Ultimately, the primary molecular roles and biological pathways of PCa were identified using enrichment analysis by the Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO). An effect of AC016773.1 on prostate cancer cell proliferation was investigated by knocking out AC016773.1 in an animal model. The interrelationship between AC016773.1 and hsa-mir-25 was validated using RNA immunoprecipitation technology. Results. The lncRNA, miRNA, and mRNA expression matrices obtained from the TCGA database contain 16901, 1881, and 19962 transcripts, respectively. Through differential expression analysis, we obtained 2010 de lncRNA comparative information, 75 lncRNA and miRNA interaction pairs, and 31 targeted de mRNAs. Through the differential expression analysis of RNA nodes in the ceRNA regulatory network, we found that compared with the NP group, in the PCa group, there were 14 lncRNAs upregulated and 25 lncRNAs downregulated, 1 miRNA upregulated and 3 miRNA downregulated, and 10 mRNA upregulated and 21 mRNA downregulated. In KEGG enrichment analysis, the pathways identified by targeted DE-mRNAs were mainly related to calcium signaling pathway, EGFR tyrosine kinase inhibitor resistance, melanoma, PI3K−Akt signaling pathway, and proteoglycans in cancer. In animal models, it was found that knocking down AC016773.1 significantly reduced tumor volume and weight, indicating that AC016773.1 may promote the proliferation of PCa cells. The use of RNA immunoprecipitation technology indicates a direct binding between AC016773.1 and hsa-mir-25. Conclusion. We elucidated the network regulatory relationship of lncRNA-miRNA-mRNA in PCa and further explored the key molecular functions, biological pathways, and prognostic value of targeted DE-mRNAs. [ABSTRACT FROM AUTHOR]

Simple Summary: Malignant meningiomas have high aggressiveness and recurrence rates and a poor prognosis, with no clear pharmacological treatment available. The study aims to investigate the progression mechanism of malignant meningiomas and the targets of intervention. RACK1 and CSNK2B have been shown to promote tumor progression but their roles in meningiomas are not clear. In this study, we will investigate the roles of RACK1-CSNK2B in meningiomas and search for targets to inhibit the progression of meningiomas. Higher-grade meningiomas (WHO grade II and III) are characterized by aggressive invasiveness and high postoperative recurrence rates. The prognosis remains inadequate even with adjuvant radiotherapy and currently there is no definitive pharmacological treatment strategy and target for malignant meningiomas. This study aims to unveil the mechanisms driving the malignant progression of meningiomas and to identify potential inhibitory targets, with significant clinical implications. Implementing techniques such as protein immunoprecipitation, mass spectrometry, RNA interference, and transcriptome sequencing, we investigated the malignancy mechanisms in meningioma cell lines IOMM-LEE and CH157-MN. Additionally, in vivo experiments were carried out on nude mice. We discovered a positive correlation between meningioma malignancy and the levels of the receptor for activated C kinase 1 (RACK1), which interacts with CSNK2B, the β subunit of casein kinase 2 (CK2), inhibiting its ubiquitination and subsequent degradation. This inhibition allows CK2 to activate the NF-κb pathway, which increases the transcription of CDK4 and cyclin D3, resulting in the transition of the cell cycle into the G2/M phase. The RACK1 inhibitor, harringtonolide (HA), significantly suppressed the malignant tendencies of meningioma cells. Our study suggests that RACK1 may play a role in the malignant progression of meningiomas, and therefore, targeting RACK1 could emerge as an effective strategy for reducing the malignancy of these tumors. [ABSTRACT FROM AUTHOR]

Simple Summary: We aimed to investigate the intrinsic biomechanical properties of melanoma cancer cells. We demonstrate that the modulation of stiffness properties, and their correlated morphometrics, impacts cutaneous melanoma cell invasiveness following EMT switching and Tspan8 transcriptional activation. Thus, we propose that melanoma stiffness measurement could be of prognostic relevance for patients. The intrinsic biomechanical properties of cancer cells remain poorly understood. To decipher whether cell stiffness modulation could increase melanoma cells' invasive capacity, we performed both in vitro and in vivo experiments exploring cell stiffness by atomic force microscopy (AFM). We correlated stiffness properties with cell morphology adaptation and the molecular mechanisms underlying epithelial-to-mesenchymal (EMT)-like phenotype switching. We found that melanoma cell stiffness reduction was systematically associated with the acquisition of invasive properties in cutaneous melanoma cell lines, human skin reconstructs, and Medaka fish developing spontaneous MAP-kinase-induced melanomas. We observed a systematic correlation of stiffness modulation with cell morphological changes towards mesenchymal characteristic gains. We accordingly found that inducing melanoma EMT switching by overexpressing the ZEB1 transcription factor, a major regulator of melanoma cell plasticity, was sufficient to decrease cell stiffness and transcriptionally induce tetraspanin-8-mediated dermal invasion. Moreover, ZEB1 expression correlated with Tspan8 expression in patient melanoma lesions. Our data suggest that intrinsic cell stiffness could be a highly relevant marker for human cutaneous melanoma development. [ABSTRACT FROM AUTHOR]

Simple Summary: Lung cancer accounts for 11.4% of all cancer cases and 18.0% of all cancer deaths. Understanding the molecular mechanisms underlying lung adenocarcinoma (LUAD), one of the most common lung cancers, is imperative to enhance diagnostic accuracy, drug development, and prognostic evaluation. The present study explored the interaction between heparan sulfate (glucosamine) 3-O-sulfotransferase 1 (HS3ST1) and Glypican 4 (GPC4), as well as the influence of the hypoxia-derived exosomal lncRNA OIP5-AS1 on the glycolysis, proliferation, and metastasis ability in LUAD cell lines and their effects on the tumor growth in xenograft animal models. GPC4 promotes HS3ST1-mediated glycolysis, and the hypoxia-derived exosomal lncRNA OIP5-AS1 promotes glycolysis via the miR-200c-3p axis in LUAD cells. The hypoxia-derived exosomal lncRNA OIP5-AS1 enhances LUAD cell proliferation and metastasis in vitro and promotes LUAD tumor growth and metastasis via miR-200c-3p in vivo. These findings highlight that the hypoxia-derived exosomal lncRNA OIP5-AS1 may participate in LUAD progression. Objective: The diagnosis of lung adenocarcinoma (LUAD) is often delayed due to the typically asymptomatic nature of the early-stage disease, causing advanced-stage LUAD diagnosis in most patients. Hypoxia is widely recognized as a driving force in cancer progression. Exosomes originating from hypoxic tumor cells promote tumorigenesis by influencing glycolysis, migration, invasion, and immune infiltration. Given these insights, our study aimed to explore the role of hypoxia-derived exosomal long non-coding RNA (lncRNA) OIP5-AS1 in LUAD cell lines and mouse models. Materials and Methods: Exosomes were meticulously isolated and authenticated based on their morphology and biomarkers. The interaction between heparan sulfate (glucosamine) 3-O-sulfotransferase 1 (HS3ST1) and Glypican 4 (GPC4) was examined using immunoprecipitation. The influence of the hypoxia-derived exosomal lncRNA OIP5-AS1 on glycolysis was assessed in LUAD cell lines. The effect of the hypoxia-derived exosomal lncRNA OIP5-AS1 on cell proliferation and metastasis was evaluated using colony formation, cell viability, cell cycle, and apoptosis analyses. Its effects on tumor size were confirmed in xenograft animal models. Results: Our study revealed the mechanism of the hypoxia-derived exosomal lncRNA OIP5-AS1 in LUAD progression. We discovered that GPC4 promotes HS3ST1-mediated glycolysis and that the hypoxia-derived exosomal lncRNA OIP5-AS1 enhances glycolysis by regulating miR-200c-3p in LUAD cells. Notably, this lncRNA stimulates LUAD cell proliferation and metastasis and fosters LUAD tumor size via miR-200c-3p. Our findings underscore the potential role of the hypoxia-derived exosomal lncRNA OIP5-AS1 in LUAD progression. Conclusions: The hypoxia-derived exosomal lncRNA OIP5-AS1 promotes LUAD by regulating HS3ST1-GPC4-mediated glycolysis via miR-200c-3p. [ABSTRACT FROM AUTHOR]

Objective. Fibroblast-like synoviocytes (FLSs) are critical for promoting joint damage in rheumatoid arthritis (RA). N6 -methyladenosine (m6 A) modification plays key roles in various diseases, but its role in the pathogenesis of RA is largely unknown. Here, we investigate increased demethylase ALKBH5 promotion of proliferation, migration, and invasion of RA FLSs via regulating JARID2 expression. Methods. ALKBH5 expression in FLSs was evaluated using real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot. 5-ethynyl-20-deoxyuridine, scratch wound healing, and transwell assays were implemented to determine the role of ALKBH5 on RA FLS proliferation, mobility, and migration. Then, m6 A sequencing combined with RNA sequencing was performed to identify the potential targets of ALKBH5. RNA immunoprecipitation and RNA pulldown were then used to validate the interaction between the protein and messenger RNA (mRNA). Collagen-induced arthritis (CIA) and delayed-type hypersensitivity arthritis (DTHA) models were further established to assess the therapeutic potency of ALKBH5 in vivo. Results. We demonstrated that ALKBH5 expression was increased in FLSs and synovium from RA. Functionally, ALKBH5 knockdown inhibited the proliferation, migration, and invasion of RA FLSs, whereas overexpression of ALKBH5 displayed the opposite effect. Mechanistically, ALKBH5 mediated m6 A modification in the JARID2 mRNA and enhanced its mRNA stability in cooperation with IGF2BP3. Intriguingly, the severity of arthritis was attenuated in mice with DTHA and ALKBH5 knockout or rats with CIA and intra-articular injection of ALKBH5 short hairpin RNA. Conclusion. Our findings suggest that ALKBH5-mediated m6 A modification is crucial for synovial hyperplasia and invasion in RA. ALKBH5 might be a potential therapeutic target for RA and even for dysregulated fibroblasts in a wide range of diseases. [ABSTRACT FROM AUTHOR]

The stimulator of interferon genes (STING) signaling pathway is crucial for the pathogenesis of autoimmune and inflammatory disorders, including acute lung injury (ALI). Apigenin (4 ′ ,5,7-trihydroxyflavone) is a natural flavonoid widely found in fruits, vegetables, and Chinese medicinal herbs that exhibits a range of pharmacological effects, such as antibacterial and anti-inflammatory activities. However, the efficacy of apigenin in STING pathway-mediated diseases remains unclear. Accordingly, this study screened Chinese medicines to identify potent agents that reduced the synthesis of type I interferons (IFNs). The results revealed apigenin as a potent compound with low cytotoxicity that markedly reduced the synthesis of type I IFNs in response to STING pathway agonists. Besides, apigenin markedly suppressed innate immune responses triggered by the STING agonist SR-717. Mechanistically, apigenin downregulated IFN beta 1 (IFNB1) expression mediated by the STING pathway via dose-dependent inhibition of STING expression, reduction of dimerization, nuclear translocation of phosphorylated IRF3, and disruption of the association between STING and IRF3. Moreover, apigenin effectively mitigated pathological pulmonary inflammation and lung edema in lipopolysaccharide (LPS)-induced ALI in mice. Apigenin further strongly attenuated the hallmarks of immoderate inflammation (interleukin (IL)-6, IL-1 β , and tumor necrosis factor α) and innate immune responses (IFNB1, C-X-C motif chemokine ligand 10, and IFN-stimulated gene 15) by preventing the activation of the STING/IRF3 pathway both in vitro and in vivo. Importantly, SR-717 significantly reversed the inhibitory effects of apigenin in LPS-induced THP1-BlueTM ISG macrophages. Collectively, apigenin effectively alleviated innate immune responses and mitigated inflammation in LPS-induced ALI via inhibition of the STING/IRF3 pathway. These findings suggest the potential of apigenin as a prophylactic and therapeutic candidate for managing STING-mediated diseases. [ABSTRACT FROM AUTHOR]

Aim: To investigate the relationship between interleukin‐17 (IL‐17), ferroptosis and osteogenic differentiation. Materials and Methods: We first analysed the changes in ferroptosis‐related molecules in experimental periodontitis models. The effects of erastin, a small‐molecule ferroptosis inducer, and IL‐17 on alveolar bone loss and repair in animal models were then investigated. Primary mouse mandibular osteoblasts were exposed to erastin and IL‐17 in vitro. Ferroptosis‐ and osteogenesis‐related genes and proteins were detected. Further, siRNA, immunofluorescence co‐localization and immunoprecipitation were used to confirm the roles of the nuclear factor erythroid‐2‐related factor 2 (NRF2) and phosphorylated signal transducer and activator of transcription 3 (p‐STAT3), as well as their interaction. Results: The levels of NRF2, glutathione peroxidase 4 and solute carrier family 7 member 11 were lower in the ligated tissues than in normal periodontal tissues. Alveolar bone loss in an in vivo experimental periodontitis model was aggravated by erastin and alleviated by IL‐17. In vitro, IL‐17 ameliorated erastin‐inhibited osteogenic differentiation by reversing ferroptosis. Altered NRF2 expression correlated with changes in ferroptosis‐related molecules and osteogenesis. Furthermore, the physical interaction between NRF2 and p‐STAT3 was confirmed in the nucleus. In IL‐17 + erastin‐stimulated osteoblasts, the p‐STAT3–NRF2 complex might actively participate in the downstream transcription of ferroptosis‐ and osteogenesis‐related genes. Conclusions: IL‐17 administration conferred resistance to erastin‐induced osteoblast ferroptosis and osteogenesis. The possible mechanism may involve p‐STAT3 directly interacting with NRF2. [ABSTRACT FROM AUTHOR]

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Background: Given that autoantibodies to peptidylarginine deiminase 4 (PAD4) are associated with erosive disease in established rheumatoid arthritis (RA), this study was conducted to compare the clinical and prognostic use of anti‐PAD4 antibodies in patients with early and established RA. Methods: Sera from patients with early (duration <2 years; n = 422) or established (duration ≥2 years; n = 359) RA from two randomized clinical trials of tofacitinib ± methotrexate compared with adalimumab + MTX or MTX alone were evaluated for the presence of anti‐PAD4 and anti‐PAD3/4 antibodies at baseline and posttreatment time points. Summary statistics were calculated for demographic, clinical, and serological characteristics, and generalized estimating equations were used to model clinical outcomes by disease duration according to anti‐PAD4 status. Results: Anti‐PAD4 antibodies were present in 22% and 40% of patients with early and established RA, respectively, stable following treatment, and associated with baseline joint damage only in established RA. In early RA, baseline anti‐PAD4 antibodies were associated with a greater improvement in disease activity score 28‐joint count using C‐reactive protein levels after treatment compared with individuals with negative anti‐PAD4 (P = 0.049). Tofacitinib ± MTX was more broadly efficacious than MTX alone at improving clinical outcomes in early and established RA, irrespective of anti‐PAD4 status (P < 0.05 for all), whereas adalimumab + MTX exhibited differential benefits in achieving disease activity score remission in early RA (P = 0.036) and American College of Rheumatology 20 responses in established RA (P = 0.002). Conclusion: Differences in prevalence, clinical associations, and treatment‐response outcomes according to anti‐PAD4 antibody status in early and established RA suggests the existence of a therapeutic window to prevent the accumulation of irreversible joint damage in early patients with RA with anti‐PAD4 antibodies. [ABSTRACT FROM AUTHOR]

Choline plays a crucial role in hepatic lipid homeostasis by acting as a major methyl-group donor. However, despite this well-accepted fact, no study has yet explored how choline's methyl-donor function contributes to preventing hepatic lipid dysregulation. Moreover, the potential regulatory role of Ire-1 α , an ER-transmembrane transducer for the unfolded protein response (UPRer), in choline-mediated hepatic lipid homeostasis remains unexplored. Thus, this study investigated the mechanism by which choline prevents hepatic lipid dysregulation, focusing on its role as a methyl-donor and the involvement of Ire-1 α in this process. To this end, a model animal for lipid metabolism, yellow catfish (Pelteobagrus fulvidraco) were fed two different diets (adequate or deficient choline diets) in vivo for 10 weeks. The key findings of studies are as follows: 1. Dietary choline, upregulated selected lipolytic and fatty acid β -oxidation transcripts promoting hepatic lipid homeostasis. 2. Dietary choline ameliorated UPRer and prevented hepatic lipid dysregulation mainly through ire-1α signalling, not perk or atf-6α signalling. 3. Choline inhibited the transcriptional expression level of ire-1 α by activating site-specific DNA methylations in the promoter of ire-1α. 4. Choline-mediated ire-1α methylations reduced Ire-1 α /Fas interactions, thereby further inhibiting Fas activity and reducing lipid droplet deposition. These results offer a novel insight into the direct and indirect regulation of choline on lipid metabolism genes and suggests a potential crosstalk between ire-1α signalling and choline-deficiency-induced hepatic lipid dysregulation, highlighting the critical contribution of choline as a methyl-donor in maintaining hepatic lipid homeostasis. [ABSTRACT FROM AUTHOR]

Background: Tissue factor (TF) activity is stringently regulated through processes termed encryption. Post-translational modification of TF and its interactions with various protein and lipid moieties allows for a multi-step de-encryption of TF and procoagulant activation. Membrane-associated guanylate kinase-with inverted configuration (MAGI) proteins are known to regulate the localisation and activity of a number of proteins including cell-surface receptors. Methods: The interaction of TF with MAGI1 protein was examined as a means of regulating TF activity. MDA-MB-231 cell line was used which express TF and MAGI1, and respond well to protease activated receptor (PAR)2 activation. Proximity ligation assay (PLA), co-immunoprecipitation and pull-down experiments were used to examine the interaction of TF with MAGI1-3 proteins and to investigate the influence of PAR2 activation. Furthermore, by cloning and expressing the PDZ domains from MAGI1, the TF-binding domain was identified. The ability of the recombinant PDZ domains to act as competitors for MAGI1, allowing the induction of TF procoagulant and signalling activity was then examined. Results: PLA and fluorescence microscopic analysis indicated that TF predominantly associates with MAGI1 and less with MAGI2 and MAGI3 proteins. The interaction of TF with MAGI1 was also demonstrated by both co-immunoprecipitation of TF with MAGI1, and co-immunoprecipitation of MAGI1 with TF. Moreover, activation of PAR2 resulted in reduction in the association of these two proteins. Pull-down assays using TF-cytoplasmic domain peptides indicated that the phosphorylation of Ser253 within TF prevents its association with MAGI1. Additionally, the five HA-tagged PDZ domains of MAGI1 were overexpressed separately, and the putative TF-binding domain was identified as PDZ1 domain. Expression of this PDZ domain in cells significantly augmented the TF activity measured both as thrombin-generation and also TF-mediated proliferative signalling. Conclusions: Our data indicate a stabilising interaction between TF and the PDZ-1 domain of MAGI1 and demonstrate that the activation of PAR2 disrupts this interaction. The release of TF from MAGI1 appears to be an initial step in TF de-encryption, associated with increased TF-mediated procoagulant and signalling activities. This mechanism is also likely to lead to further interactions and modifications leading to further enhancement of procoagulant activity, or the release of TF. [ABSTRACT FROM AUTHOR]

Simple Summary: RNA-binding proteins (RBPs) play a critical role in controlling gene expression post-transcriptionally and their dysregulation can lead to various diseases, including cancer. One such RBP is HuR, whose levels have been associated with a poor clinical prognosis in breast cancer. Nevertheless, the precise molecular mechanism underlying this connection remains incompletely characterized. Our study uncovers that HuR targets SOX9 mRNA in breast cancer cells and provides compelling evidence supporting HuR's involvement in cell migration and invasion. RNA-binding proteins play diverse roles in cancer, influencing various facets of the disease, including proliferation, apoptosis, angiogenesis, senescence, invasion, epithelial–mesenchymal transition (EMT), and metastasis. HuR, a known RBP, is recognized for stabilizing mRNAs containing AU-rich elements (AREs), although its complete repertoire of mRNA targets remains undefined. Through a bioinformatics analysis of the gene expression profile of the Hs578T basal-like triple-negative breast cancer cell line with silenced HuR, we have identified SOX9 as a potential HuR-regulated target. SOX9 is a transcription factor involved in promoting EMT, metastasis, survival, and the maintenance of cancer stem cells (CSCs) in triple-negative breast cancer. Ribonucleoprotein immunoprecipitation assays confirm a direct interaction between HuR and SOX9 mRNA. The half-life of SOX9 mRNA and the levels of SOX9 protein decreased in cells lacking HuR. Cells silenced for HuR exhibit reduced migration and invasion compared to control cells, a phenotype similar to that described for SOX9-silenced cells. [ABSTRACT FROM AUTHOR]

Background: Usenamine A, a novel natural compound initially isolated from the lichen Usnea longissima, has exhibited promising efficacy against hepatoma in prior investigation. Nevertheless, the underlying mechanisms responsible for its antihepatoma effects remain unclear. Furthermore, the role of the AKT/mechanistic target of the rapamycin (mTOR)/signal transducer and activator of transcription 3 (STAT3)/inhibitor of differentiation/DNA binding 1 (ID1) signaling axis in hepatocellular carcinoma (HCC), and the potential anti-HCC effects of drugs targeting this pathway are not well understood. Methods: CCK-8 assay was used to investigate the effects of usenamine A on the proliferation of human HCC cells. Moreover, the effects of usenamine A on the invasion ability of human HCC cells were evaluated by transwell assay. In addition, expression profiling analysis, quantitative real-time PCR, immunoblotting, immunohistochemistry (IHC) analysis, RNAi, immunoprecipitation, and chromatin immunoprecipitation (ChIP) assay were used to explore the effects of usenamine A on the newly identified AKT/mTOR/STAT3/ID1 signaling axis in human HCC cells. Results: Usenamine A inhibited the proliferation and invasion of human HCC cell lines (HepG2 and SK-HEP-1). Through the analysis of gene expression profiling, we identified that usenamine A suppressed the expression of ID1 in human HCC cells. Furthermore, immunoprecipitation experiments revealed that usenamine A facilitated the degradation of the ID1 protein via the ubiquitin–proteasome pathway. Moreover, usenamine A inhibited the activity of STAT3 in human HCC cells. ChIP analysis demonstrated that STAT3 positively regulated ID1 expression at the transcriptional level in human HCC cells. The STAT3/ID1 axis played a role in mediating the anti-proliferative and anti-invasive impacts of usenamine A on human HCC cells. Additionally, usenamine A suppressed the STAT3/ID1 axis through AKT/mTOR signaling in human HCC cells. Conclusion: Usenamine A displayed robust anti-HCC potential, partly attributed to its capacity to downregulate the AKT/mTOR/STAT3/ID1 signaling pathway and promote ubiquitin–proteasome-mediated ID1 degradation. Usenamine A has the potential to be developed as a therapeutic agent for HCC cases characterized by abnormal AKT/mTOR/STAT3/ID1 signaling, and targeting the AKT/mTOR/STAT3 signaling pathway may be a viable option for treating patients with HCC exhibiting elevated ID1 expression. [ABSTRACT FROM AUTHOR]

Objective: Autoantibodies are clinically useful in phenotyping patients with systemic sclerosis (SSc). Gastrointestinal (GI) function is regulated by the enteric nervous system (ENS) and commonly impaired in SSc, suggesting that the SSc autoimmune response may target ENS antigens. We sought to identify novel anti‐ENS autoantibodies with an aim to clinically phenotype SSc GI dysfunction. Methods: Serum from a patient with SSc with GI dysfunction but without defined SSc‐associated autoantibodies was used for autoantibody discovery. Immunoprecipitations performed with murine myenteric plexus lysates were on‐bead digested, and autoantigens were identified by mass spectrometry. Prevalence was determined, and clinical features associated with novel autoantibodies were evaluated in a SSc cohort using regression analyses. The expression of gephyrin in human GI tract tissue was examined by immunohistochemistry. Results: We identified gephyrin as a novel SSc autoantigen. Anti‐gephyrin antibodies were present in 9% of patients with SSc (16/188) and absent in healthy controls (0/46). Anti‐gephyrin antibody–positive patients had higher constipation scores (1.00 vs 0.50, P = 0.02) and were more likely to have severe constipation and severe distention/bloating (46% vs 15%, P = 0.005; 54% vs 25%, P = 0.023, respectively). Anti‐gephyrin antibody levels were significantly higher among patients with severe constipation (0.04 vs 0.00; P = 0.001) and severe distention and bloating (0.03 vs 0.004; P = 0.010). Severe constipation was associated with anti‐gephyrin antibodies even in the adjusted model. Importantly, gephyrin was expressed in the ENS, which regulates gut motility. Conclusion: Gephyrin is a novel ENS autoantigen that is expressed in human myenteric ganglia. Anti‐gephyrin autoantibodies are associated with the presence and severity of constipation in patients with SSc. [ABSTRACT FROM AUTHOR]

Indirect immunofluorescence on HEp-2 cell line (HEp-2-IIF) remains "gold standard" method for the detection of antinuclear antibodies (ANA). ANA is an operative definition, showing the possibility of autoantibodies (Aab) to bind nuclear, and cytoplasmic antigens. One of the major examples is represented by anti-mitochondrial antibodies (AMAs), which target proteins of the inner and outer mitochondrial membranes, located into the cytoplasm. The standard IIF on rat kidney/stomach/liver tissue sections, with the combined use of other commercial assays, may all be used in ordinary lab life to validate the AC-21 pattern on Hep-2 cells. The routine lab experience teaches that commercial kits cannot always be detected and define specific AMAs. In these cases the literature proposes the use of other homemade assays to detect AMAs as immunoprecipitation (IP) and Western blot (IP-WB). However, using IP or IP-WB is difficult to apply in a routine laboratory, because of numerous cases to process and the related troubles. Where find confirmation of the AC-21 pattern if line-immunoblot and other routine methods (ELISA, CLIA/FEIA assays) fail? We review AC-21 AMA-like sera from our patients (year 2022) and propose a revised diagnostic algorithm based on the combined use of IIF on Hep-2 cells, line immunoblot and IIF on rodent tissue as a third line method. We demonstrated that, particularly in cases where the second level test was unsuccessful, the application of IFI on rodent tissues became indispensable to verify the existence of AMAs. [ABSTRACT FROM AUTHOR]

Introduction. Endometriosis (EMs), manifested by pain and infertility, is a chronic inflammatory disease. The precise pathophysiology of this disease remains uncertain. Insulin-like growth factor-2 mRNA-binding protein 1 (IGF2BP1) and polypyrimidine tract-binding protein 1 (PTBP1) have both been found to regulate proliferation, apoptosis, and invasion. This study aimed to investigate the effects of IGF2BP1/PTBP1 in treating EMs. Materials and methods. qRT-PCR and western blotting were employed to quantify IGF2BP1 and PTBP1 expression in six patients with EMs (mean age 33.83 years). The correlation analysis, STRING database prediction, and RNA immunoprecipitation were utilized to identify the relationship between IGF2BP1 and PTBP1. Ectopic endometrial volume, weight, HE staining, and IGF2BP1 silencing were utilized to estimate the effects of IGF2BP1 in EMs model rats. qRT-PCR, CCK-8, 5-ethynyl-2’-deoxyuridine (EDU) labeling, Transwell assay, and flow cytometry were utilized to assess the effects of IGF2BP1/PTBP1 on the proliferation, migration, invasion, and apoptosis of ectopic endometrial stromal cells (eESCs). Furthermore, western blotting was employed to evaluate expressions of PCNA, VEGF, and E-cadherin in EMs rats and eESCs. Results. The mRNA and protein levels of IGF2BP1 and PTBP1 in the ectopic and eutopic endometrium of EMs patients were significantly increased. RNA immunoprecipitation revealed a close interaction of IGF2BP1 with PTBP1. Additionally, the endometrial volume, weight, and histopathologic scores in rats were significantly reduced after IGF2BP1 silencing. IGF2BP1 silencing also decreased the expression of PCNA and VEGF, and increased E-cadherin expression in endometrial tissues of EMs rats. Moreover, IGF2BP1 silencing inhibited proliferation, migration, and invasion and promoted apoptosis through PTBP1 in eESCs. Conclusions. IGF2BP1 exhibits potential beneficial properties in the management of EMs by interacting with PTBP1, thereby highlighting IGF2BP1 as a promising therapeutic target for EMs. [ABSTRACT FROM AUTHOR]

Introduction: Alzheimer's disease (AD) is the most prevalent neurodegenerative disease characterized by extracellular senile plaques including amyloid-β peptides and intracellular neurofibrillary tangles consisting of abnormal Tau. Depression is one of the most common neuropsychiatric symptoms in AD, and clinical evidence demonstrates that depressive symptoms accelerate the cognitive deficit of AD patients. However, the underlying molecular mechanisms of depressive symptoms present in the process of AD remain unclear. Methods: Depressive-like behaviors and cognitive decline in hTau mice were induced by chronic restraint stress (CRS). Computational prediction and molecular experiments supported that an asparagine endopeptidase (AEP)- derived Tau fragment, Tau N368 interacts with peroxisome proliferator-activated receptor delta (PPAR-δ). Further behavioral studies investigated the role of Tau N368-PPAR-δ interaction in depressive-like behaviors and cognitive declines of AD models exposed to CRS. Results: We found that mitochondrial dysfunction was positively associated with depressive-like behaviors and cognitive deficits in hTau mice. Chronic stress increased Tau N368 and promoted the interaction of Tau N368 with PPAR-δ, repressing PPAR-δ-mediated transactivation in the hippocampus of mice. Then we predicted and identified the binding sites of PPAR-δ. Finally, inhibition of AEP, clearance of Tau N368 and pharmacological activation of PPAR-δ effectively alleviated CRS-induced depressive-like behaviors and cognitive decline in mice. Conclusion: These results demonstrate that Tau N368 in the hippocampus impairs mitochondrial function by suppressing PPAR-δ, facilitating the occurrence of depressive-like behaviors and cognitive decline. Therefore, our findings may provide new mechanistic insight in the pathophysiology of depression-like phenotype in mouse models of Alzheimer's disease. [ABSTRACT FROM AUTHOR]

Simple Summary: Head and neck squamous cell carcinoma (HNSCC) ranks seventh among malignant tumors worldwide, with an estimated 500,000 new cases annually. Despite the development of early diagnosis technology, there are still 60% of patients already in the middle–late stages when they were first diagnosed with HNSCC. Despite advances in multidisciplinary synthetic therapy, the overall survival rate remains low, with a five-year survival rate of less than 50%. Thus, it is urgently needed to search for novel early diagnosis biomarkers and therapy targets. In this study, we revealed the role of SOX2-OT in HNSCC progression and metastasis by binding with ILF3, which may serve as a therapeutic target and prognostic biomarker to improve the early diagnosis and overall survival of HNSCC. Long non-coding RNA (lncRNA) is involved in the progression of head and neck squamous cell carcinoma (HNSCC). The molecular mechanism of lncRNA SOX2-OT in HNSCC remains unclear. Therefore, we aimed to elucidate the oncogenic role of SOX2-OT in HNSCC. QRT-PCR analysis was performed in 61 pairs of HNSCC cancer tissues, adjacent normal tissues, and 68 plasma samples confirmed that lncRNA SOX2-OT was overexpressed in cancer tissues and plasma samples, which served as a poor prognostic factor for HNSCC. The FISH assay demonstrated that SOX2-OT was localized in the nucleus and cytoplasm of HNSCC cell lines. Further, the cell function assay confirmed that SOX2-OT promoted cell proliferation and metastasis in vitro and in vivo. RNA pulldown and RIP assay results revealed that SOX2-OT bonds with ILF3 in HNSCC, and the rescue assay confirmed that SOX2-OT played an oncogenic role depending on ILF3 protein expression. Ingenuity pathway analysis and Western blotting indicated that SOX2-OT regulated HNSCC progression by promoting STAT3 phosphorylation and modulating the crosstalk between STAT3 and TGF-β signaling. These results reveal evidence for the role of SOX2-OT in HNSCC progression and metastasis by binding to ILF3, which may serve as a therapeutic target and prognostic biomarker in HNSCC. [ABSTRACT FROM AUTHOR]

Simple Summary: Emerging evidence suggests that AR is involved in the growth of esophageal squamous cell carcinoma. However, how AR exerts its functions in the invasion of esophageal squamous cell carcinoma remains unknown. In this study, by analyzing our previous AR cistromes and androgen-regulated transcriptomes, we find that AR upregulates the expression of UGT2B15 in esophageal cancer cells. We further find that the transcription factor AP-1 governs AR-induced transcription. Importantly, genetic or pharmacological inhibition of AP-1 relieves AR-mediated UGT2B15 activation, leading to esophageal cancer cell invasion inhibition. Our findings reveal the molecular mechanisms of AR in esophageal cancer invasion and identify the AP1-directed AR transcriptional activation of UGT2B15 as a potential therapeutic target for esophageal cancer metastasis. Esophageal squamous cell carcinoma (ESCC) is an aggressive epithelial malignancy with poor prognosis. Interestingly, ESCC is strongly characterized by a male-predominant propensity. Our previous study showed that androgen receptor (AR) orchestrated a transcriptional repression program to promote ESCC growth, but it remains unclear whether AR can also activate oncogenic signaling during ESCC progression. In this study, by analyzing our previous AR cistromes and androgen-regulated transcriptomes, we identified uridine diphosphate glucuronosyltransferase family 2 member B15 (UGT2B15) as a bona fide target gene of AR. Mechanistically, AP-1 cofactors played important and collaborative roles in AR-mediated UGT2B15 upregulation. Functional studies have revealed that UGT2B15 promoted invasiveness in vitro and lymph node metastasis in vivo. UGT2B15 was partially responsible for the AR-induced invasive phenotype in ESCC cells. Importantly, simultaneous blocking of AP-1 and AR resulted in stronger inhibition of cell invasiveness compared to inhibiting AP-1 or AR alone. In conclusion, our study reveals the molecular mechanisms underlying the AR-driven ESCC invasion and suggests that the AR/AP1/UGT2B15 transcriptional axis can be potentially targeted in suppressing metastasis in male ESCC patients. [ABSTRACT FROM AUTHOR]

Background: Circular RNAs (circRNAs) are involved in the malignant development of tumors. However, the mechanism of circ_0102231 in non‐small cell lung cancer (NSCLC) has rarely been discussed and reported. Methods: Quantitative real‐time polymerase chain reaction (qRT‐PCR) was used to measure the expression of circ_0102231, miR‐635 and NOVA alternative splicing regulator 2 (NOVA2) in NSCLC tissues and cells. Western blot was applied to detect the protein expression. Cell proliferation was monitored by cell counting kit‐8 (CCK8) and 5‐ethynyl‐2′‐deoxyuridine (EdU) experiments. The angiogenesis ability of cells was tested by angiogenesis assay. Flow cytometry was used to analyze cell apoptosis. The relationship between circ_0102231 and NOVA2 or miR‐635 was analyzed by dual‐luciferase reporter assay and RNA immunoprecipitation (RIP) assay. An in vivo transplanted tumor model was established to confirm the effect of circ_0102231 on tumor formation. Results: Circ_0102231 was abnormally upregulated in NSCLC tissues and correlated with clinical stage. Silencing of circ_0102231 inhibited cell proliferation and angiogenesis but significantly promoted the apoptosis of NSCLC cells. There were target binding sites between circ_0102231 and miR‐635, miR‐635 and NOVA2. Importantly, circ_0102231 acted as a sponge for miR‐635, increased the expression of NOVA2, and activated the PI3K/AKT signaling pathway. Finally, silencing of circ_0102231 also had obvious antitumor effects in vivo. Conclusion: Circ_0102231 increased the expression of NOVA2 by interacting with miR‐635 to promote the malignant progression of NSCLC. [ABSTRACT FROM AUTHOR]

Background: Salvianolic acid B (Sal B), a water-soluble phenolic compound derived from Salvia miltiorrhiza Bunge, is commonly used in Traditional Chinese Medicine to treat cardiovascular disease. In our previous study, Sal B protected against myocardial fibrosis induced by diabetic cardiomyopathy (DCM). This study aimed to investigate the ameliorative effects and potential mechanisms of Sal B in mitigating myocardial fibrosis induced by DCM. Methods: Various methods were used to investigate the effects of Sal B on myocardial fibrosis induced by DCM in vivo and in vitro. These methods included blood glucose measurement, echocardiography, HE staining, Masson's trichrome staining, Sirius red staining, cell proliferation assessment, determination of hydroxyproline levels, immunohistochemical staining, evaluation of fibrosis-related protein expression (Collagen-I, Collagen-III, TGF-β1, p-Smad3, Smad3, Smad7, and α-smooth muscle actin), analysis of Smad7 gene expression, and analysis of Smad7 ubiquitin modification. Results: The animal test results indicated that Sal B significantly improved cardiac function, inhibited collagen deposition and phenotypic transformation, and ameliorated myocardial fibrosis in DCM by upregulating Smad7, thereby inhibiting the TGF-β1 signaling pathway. In addition, cell experiments demonstrated that Sal B significantly inhibited the proliferation, migration, phenotypic transformation, and collagen secretion of cardiac fibroblasts (CFs) induced by high glucose (HG). Sal B significantly decreased the ubiquitination of Smad7 and stabilized the protein expression of Smad7, thereby increasing the protein expression of Smad7 in CFs and inhibiting the TGF-β1 signaling pathway, which may be the potential mechanism by which Sal B mitigates myocardial fibrosis induced by DCM. Conclusion: This study revealed that Sal B can improve myocardial fibrosis in DCM by deubiquitinating Smad7, stabilizing the protein expression of Smad7, and blocking the TGF-β1 signaling pathway. [ABSTRACT FROM AUTHOR]

Objective: The aim of the study was to investigate the role of N6‐methyladenosine (m6A) modification in the progression of rheumatoid arthritis (RA). Methods: Peripheral blood mononuclear cells (PBMCs) from patients with RA and healthy controls were collected. The expression of m6A modification‐related proteins and m6A levels were detected using polymerase chain reaction (PCR), western blot, and m6A enzyme‐linked immunosorbent assay (ELISA). The roles of methyltransferase‐like 14 (METTL14) in the regulation of inflammation in RA was explored using methylated RNA immunoprecipitation (MeRIP) sequencing and RNA immunoprecipitation assays. Collagen antibody‐induced arthritis (CAIA) mice were used as an in vivo model to study the role of METTL14 in the inflammation progression of RA. Results: We found that m6A writer METTL14 and m6A levels were decreased in PBMCs of patients with active RA and correlated negatively with the disease activity score using 28 joint counts (DAS28). Knockdown of METTL14 downregulated m6A and promoted the secretion of inflammatory cytokines interleukin 6 (IL‐6) and IL‐17 in PBMCs of patients with RA. Consistently, METTL14 knockdown promoted joint inflammation accompanied by upregulation of IL‐6 and IL‐17 in CAIA mice. MeRIP sequencing and functional studies confirmed that tumor necrosis factor α induced protein 3 (TNFAIP3), a key suppressor of the nuclear factor‐κB inflammatory pathway, was involved in m6A‐regulated PBMCs. Mechanistic investigations revealed that m6A affected TNFAIP3 expression by regulation of messenger RNA stability and translocation in TNFAIP3 protein coding sequence. Conclusions: Our study highlights the critical roles of m6A on regulation of inflammation in RA progression. Treatment strategies targeting m6A modification may represent a new option for management of RA. [ABSTRACT FROM AUTHOR]

Simple Summary: Gastric cancer (GC) is one of the most common malignancies with limited therapeutic targets available. CDR1as, an endogenous circular RNA with a closed loop structure, has been reported to be a crucial regulator and promising biomarker in various tumors. However, whether and how CDR1as participates in GC progression remains not well characterized. In this study, we explored the biological roles, the downstream molecular mechanism and the upstream regulator of CDR1as in GC. We found that CDR1as mediated by human antigen R (HuR) promotes GC growth through the miR-299-3p/TGIF1 axis, which provides new insights into GC pathogenesis and brings a new potential target for clinical GC therapy. Background: Gastric cancer (GC) remains a common malignancy worldwide with a limited understanding of the disease mechanisms. A novel circular RNA CDR1as has been recently reported to be a crucial regulator of human cancer. However, its biological role and mechanism in the GC growth are still far from clear. Methods: Small interfering RNAs (siRNAs), lentivirus or plasmid vectors were applied for gene manipulation. The CDR1as effects on the GC growth were evaluated in CCK8 and colony formation assays, a flow cytometry analysis and mouse xenograft tumor models. A bioinformatics analysis combined with RNA immunoprecipitation (RIP), RNA pull-down assays, dual-luciferase reporter gene assays, Western blot, reverse transcription–quantitative polymerase chain reaction (RT-qPCR) and functional rescue experiments were used to identify the CDR1as target miRNA, the downstream target gene and its interaction with human antigen R (HuR). Results: The CDR1as overexpression promoted the GC growth in vitro and in vivo and reduced the apoptotic rate of GC cells. Its knockdown inhibited the GC cell proliferation and viability and increased the cell apoptotic rate. Proliferation-related proteins PCNA and Cyclin D1 and apoptosis-related proteins Bax, Bcl-2, Caspase-3 and Caspase-9 were regulated. Mechanically, the cytoplasmic CDR1as acted as a miR-299-3p sponge to relieve its suppressive effects on the GC cell growth. Oncogenic TGIF1 was a miR-299-3p downstream target gene that mediated the promotive effects of CDR1as and regulated the PCNA and Bax levels. HuR interacted with CDR1as via the RRM2 domain and positively regulated the CDR1as level and its oncogenic role as well as downstream target TGIF1. Conclusions: CDR1as promotes the GC growth through the HuR/CDR1as/miR-299-3p/TGIF1 axis and could be used as a new therapeutic target for GC. [ABSTRACT FROM AUTHOR]

Background: Circular RNA (circRNA) can regulate the progression of hepatocellular carcinoma (HCC). However, the role and potential mechanism of circ_0004913 in HCC are not explored. Methods: Circ_0004913 was identified from two GSE datasets (GSE94508 and GSE97322) as a differentially expressed circRNA between HCC and normal tissues. Levels of circ_0004913, microRNA-184 (miR-184), and hepcidin (HAMP) were determined by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation, migration, and invasion were estimated by methyl thiazolyl tetrazolium, colony formation, and Transwell assays, respectively. Levels of all proteins were examined by Western blot. Glucose consumption and lactate and ATP production were analyzed by the glucose, lactate, and ATP assay kits. Dual-luciferase reporter, RNA immunoprecipitation (RIP), and RNA pull-down assays were performed to verify the interactions among miR-184 and circ_0004913 or HAMP. The mice xenograft models were established to assess the effect of circ_0004913 on tumor growth in vivo. Results: Circ_0004913 was downregulated in HCC, and its expression impeded cell proliferation, migration, and invasion, EMT, and glycolysis in HCC cells. miR-184 was identified as a target miRNA of circ_0004913, and their expression levels were negatively correlated. miR-184 overexpression could reverse the inhibitory effect of circ_0004913 on HCC cell progression. Moreover, as a target gene of miR-184, HAMP expression was positively correlated with circ_0004913 expression in HCC tissues, and repression of miR-184 could inhibit the progression of HCC cells by increasing HAMP expression. Circ_0004913 could inhibit JAK2/STAT3/AKT signaling pathway and tumor growth in vivo by regulating the miR-184/HAMP axis. Conclusion: Circ_0004913 inhibited the tumorigenesis of HCC by sponging miR-184 to regulate HAMP expression in vitro and in vivo. [ABSTRACT FROM AUTHOR]

Background: Hepatocellular carcinoma (HCC) is the most common form of liver cancer. Circular RNAs (circRNAs) play a vital role in cancer development and progression. This study investigated the role and potential mechanism of circRNA filamin binding LIM protein 1 (circFBLIM1) in HCC. Methods: Exosomes were identified by transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blot assay. The levels of circFBLIM1, miR-338, and low-density lipoprotein receptor-related protein 6 (LRP6) were measured by quantitative real-time polymerase chain reaction or Western blot. Glycolysis was analyzed by detecting glucose consumption, lactate production, ATP level, extracellular acidification rate (ECAR), and oxygen consumption rate (OCR). Cell viability was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay. Cell apoptosis was detected by flow cytometry. Xenograft assay was performed to analyze tumor growth in vivo. The interaction among circFBLIM1, miR-338, and LRP6 was confirmed by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. This study was approved by the Institutional Review Board of the First Affiliated Hospital of Guizhou University of Traditional Chinese Medicine. Results: CircFBLIM1 was highly expressed in HCC serum exosomes and HCC cells. Inhibition of circFBLIM1 confined HCC glycolysis and progression. CircFBLIM1 knockdown blocked tumorigenesis in vivo. CircFBLIM1 was a sponge of miR-338 and promoted HCC progression and glycolysis by regulating miR-338. Moreover, miR-338 suppressed HCC progression and glycolysis via targeting LRP6. Mechanistically, circFBLIM1 functioned as an miR-338 sponge to upregulate LRP6. Conclusion: CircFBLIM1 facilitated HCC progression and glycolysis via modulating the miR-338/LRP6 axis, which may provide promising therapeutic targets for HCC. [ABSTRACT FROM AUTHOR]

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Objective: This research aimed to elucidate the molecular mechanisms underlying the periodontitis‐associated bone loss, with particular emphasis on the contributory role of hypoxic microenvironment in this process. Background: Periodontitis generally causes alveolar bone loss and is often associated with a hypoxic microenvironment, which affects bone homeostasis. However, the regulating mechanism between hypoxia and jaw metabolism remains unclear. Hypoxia triggers autophagy, which is closely related to osteogenic differentiation, but how hypoxia‐induced autophagy regulates bone metabolism is unknown. HDAC6 and FOXO1 are closely related to bone metabolism and autophagy, respectively, but whether they are related to hypoxia‐induced bone loss and their internal mechanisms is still unclear. Methods: Established rat nasal obstruction model and hypoxia cell model. Immunohistochemistry was performed to detect the expression and localization of HDAC6 and FOXO1 proteins, analysis of autophagic flux and transmission electron microscopy was used to examine the autophagy level and observe the autophagosomes, co‐immunoprecipitation and chromatin immunoprecipitation were preformed to investigate the interaction of HDAC6 and FOXO1. Results: Hypoxia causes increased autophagy and reduced osteogenic differentiation in rat mandibles and BMSCs, and blocking autophagy can attenuate hypoxia‐induced osteogenic differentiation decrease. Moreover, hypoxia dissociated the FOXO1‐HDAC6 complex and accumulated them in the nucleus. Knocking down of FOXO1 or HDAC6 alleviated hypoxia‐induced autophagy elevation or osteogenic differentiation reduction by binding to related genes, respectively. Conclusion: Hypoxia causes mandibular bone loss and autophagy elevation. Mechanically, hypoxia dissociates the FOXO1‐HDAC6 complex and aggregates them in the nucleus, whereas HDAC6 decreases osteogenic differentiation and FOXO1 enhances autophagy to inhibit osteogenic differentiation. [ABSTRACT FROM AUTHOR]

Background: Human bone marrow mesenchymal stem cells (hBMSCs) are a major source of osteoblast precursor cells and are directly involved in osteoporosis (OP) progression. Bromodomain-containing protein 4 (BRD4) is an important regulator for osteogenic differentiation. Therefore, its role and mechanism in osteogenic differentiation process deserve further investigation. Methods: hBMSCs osteogenic differentiation was evaluated by flow cytometry, alkaline phosphatase assay and alizarin red staining. Western blot was used to test osteogenic differentiation-related proteins, BRD4 protein, WNT family members-4 (WNT4)/NF-κB-related proteins, and glycolysis-related proteins. Metabolomics techniques were used to detect metabolite changes and metabolic pathways. BRD4 and WNT4 mRNA levels were determined using quantitative real-time PCR. Dual-luciferase reporter assay and chromatin immunoprecipitation assay were performed to detect BRD4 and WNT4 interaction. Glycolysis ability was assessed by testing glucose uptake, lactic acid production, and ATP levels. Results: After successful induction of osteogenic differentiation, the expression of BRD4 was increased significantly. BRD4 knockdown inhibited hBMSCs osteogenic differentiation. Metabolomics analysis showed that BRD4 expression was related to glucose metabolism in osteogenic differentiation. Moreover, BRD4 could directly bind to the promoter of the WNT4 gene. Further experiments confirmed that recombinant WNT4 reversed the inhibition effect of BRD4 knockdown on glycolysis, and NF-κB inhibitors (Bardoxolone Methyl) overturned the suppressive effect of BRD4 knockdown on hBMSCs osteogenic differentiation. Conclusion: BRD4 promoted hBMSCs osteogenic differentiation by inhibiting NF-κB pathway via enhancing WNT4 expression. [ABSTRACT FROM AUTHOR]

Background: Osteoarthritis (OA) is a chronic and degenerative bone and joint disease, and paeoniflorin shows anti-arthritis role in OA. This study planned to investigate the mechanism related to chondroprotective role of paeoniflorin in OA. Methods: Real-time quantitative PCR and western blotting were performed to measure expression levels of circ-PREX1, microRNA (miR)-140-3p, Wingless-type MMTV integration site family, member 5B (WNT5B), B cell lymphoma (Bcl)-2, and Bcl-2 Associated X Protein (Bax). MTT assay, EdU assay, flow cytometry and enzyme-linked immunosorbent assay evaluated cell viability, proliferation, apoptosis and inflammatory response, respectively. Dual-luciferase reporter assay and RNA immunoprecipitation assay identified the relationship among circ-PREX1, miR-140-3p, and WNT5B. Results: IL-1β highly induced apoptosis rate, Bax expression and TNF-α product, accompanied with decreased cell viability, cell proliferation and IL-10 secretion, whereas these effects were partially reversed after paeoniflorin pretreatment. Expression of circ-PREX1 was upregulated and miR-140-3p was downregulated in cartilage tissues of patients with knee OA (KOA) and IL-1β-induced human chondrocytes (C28/I2). Circ-PREX1 overexpression and miR-140-3p silencing attenuated the suppressive effect of paeoniflorin in IL-1β-induced C28/I2 cells. Furthermore, miR-140-3p was negatively regulated by circ-PREX1. WNT5B was a downstream target of miR-140-3p and could be modulated by the circ-PREX1/miR-140-3p pathway in IL-1β-induced C28/I2 cells. Conclusion: Paeoniflorin might protect human chondrocytes from IL-1β-induced inflammatory injury via circ-PREX1-miR-140-3p-WNT5B pathway, suggesting a potential preventative agent and a novel target for the treatment of KOA. [ABSTRACT FROM AUTHOR]

Simple Summary: This study explores the role of circTPST2, a circular RNA, in the chemotherapy sensitivity of head and neck squamous cell carcinoma (HNSCC). Elevated levels of circTPST2 were observed in HNSCC tissues, and functional experiments revealed its inhibitory effect on cisplatin sensitivity in HNSCC cells. By interacting with miR-770-5p and the Nucleolin pathway, circTPST2 emerged as a crucial regulator influencing chemotherapy responsiveness. The findings suggest that circTPST2 may serve as a potential marker for chemotherapy regimen selection in HNSCC patients, providing insights to enhance treatment efficacy and patient outcomes. Chemoresistance poses a significant challenge in the treatment of advanced head and neck squamous cell cancer (HNSCC). The role and mechanism of circular RNAs (circRNAs) in HNSCC chemoresistance remain understudied. We conducted circRNA microarray analysis to identify differentially expressed circRNAs in HNSCC. The expression of circRNAs from the tyrosylprotein sulfotransferase 2 (TPST2) gene and miRNAs was evaluated through qPCR, while the circular structure of circTPST2 was verified using Sanger sequencing and RNase R. Through Western blotting, biotin-labeled RNA pulldown, RNA immunoprecipitation, mass spectrometry, and rescue experiments, we discovered miR-770-5p and nucleolin as downstream targets of circTPST2. Functional tests, including CCK8 assays and flow cytometry, assessed the chemoresistance ability of circTPST2, miR-770-5p, and Nucleolin. Additionally, FISH assays determined the subcellular localization of circTPST2, miR-770-5p, and Nucleolin. IHC staining was employed to detect circTPST2 and Nucleolin expression in HNSCC patients. circTPST2 expression was inversely correlated with cisplatin sensitivity in HNSCC cell lines. Remarkably, high circTPST2 expression correlated with lower overall survival rates in chemotherapeutic HNSCC patients. Mechanistically, circTPST2 reduced chemosensitivity through sponge-like adsorption of miR-770-5p and upregulation of the downstream protein Nucleolin in HNSCC cells. The TCGA database revealed improved prognosis for patients with low circTPST2 expression after chemotherapy. Moreover, analysis of HNSCC cohorts demonstrated better prognosis for patients with low Nucleolin protein expression after chemotherapy. We unveil circTPST2 as a circRNA associated with chemoresistance in HNSCC, suggesting its potential as a marker for selecting chemotherapy regimens in HNSCC patients. Further exploration of the downstream targets of circTPST2 advanced our understanding and improved treatment strategies for HNSCC. [ABSTRACT FROM AUTHOR]

Objectives: Increasing evidence indicated circRNAs were involved in stem cells osteogenesis differentiation. Herein, we aimed to clarify the role of hsa‐circ‐0107593 during the osteogenesis process of human adipose‐derived stem cells (hADSCs) and the underlying mechanisms. Methods: The ring structure of hsa‐circ‐0107593 was confirmed using RNase R treatment and Sanger sequencing. Nucleoplasmic separation and fluorescence in situ hybridization detected hsa‐circ‐0107593 distribution. Lentivirus and siRNA were used to modulate the expression of hsa‐circ‐0107593, and the binding relationship between hsa‐circ‐0107593 and miR‐20a‐5p was verified by luciferase assay and RNA immunoprecipitation. We detected the osteogenic activity of hADSCs through alkaline phosphatase staining, alizarin red S staining, real‐time polymerase chain reaction (RT‐PCR), western blot, and cellular immunofluorescence experiment. In vivo, micro‐computed tomography was performed to analyze bone formation around skull defect. Results: RT‐PCR results exhibited that hsa‐circ‐0107593 was downregulated while miR‐20a‐5p was upregulated during hADSCs osteogenesis. In vivo and in vitro experiments results indicated that knocking down hsa‐circ‐0107593 promoted the osteogenic differentiation of hADSCs, while overexpression of hsa‐circ‐0107593 showed an inhibitory effect on hADSCs osteogenic differentiation. In vitro experiment results showed hsa‐circ‐0107593 acted as a hADSCs osteogenic differentiation negative factor for it inhibited the suppressing effect of miR‐20a‐5p on SMAD6. Conclusion: Knocking down hsa‐circ‐0107593 acts as a positive factor of the osteogenic differentiation of hADSCs via miR‐20a‐5p/SMAD6 signaling. [ABSTRACT FROM AUTHOR]

Background: Osteosarcoma (OS) is a common, aggressive primary sarcoma of bone. Drug resistance is a huge obstacle to chemotherapy for cancer. This study aimed to investigate the role and mechanism of circ_0002060 in OS resistance to doxorubicin (DOX). Methods: The levels of circ_0002060, miR-198, and ATP-binding cassette subfamily B member 1 (ABCB1) in OS tissues and DOX-resistant OS cells were measured by quantitative real-time polymerase chain reaction or Western blot assay. Kaplan–Meier analysis was performed to determine the relationship between circ_0002060 expression in OS tissues and overall survival of OS patients. The half-inhibitory concentration (IC50) of DOX was calculated using the Cell Counting Kit-8 (CCK-8) assay. Proliferation and apoptosis of DOX-resistant OS cells were assessed by colony formation assay and flow cytometry. The levels of apoptosis-related proteins in DOX-resistant OS cells were measured by Western blot assay. Xenograft assay was utilized to analyze the effect of circ_0002060 on DOX resistance in vivo. The interactions among circ_0002060, miR-198, and ABCB1 in DOX-resistant OS cells were confirmed by dual-luciferase reporter assay, RNA immunoprecipitation assay, or RNA pull-down assay. Results: circ_0002060 and ABCB1 were upregulated, while miR-198 was downregulated in OS tissues and DOX-resistant OS cells. circ_0002060 silencing reduced DOX resistance in vitro and in vivo. Moreover, circ_0002060 enhanced DOX resistance by sponging miR-198. Besides, miR-198 decreased DOX resistance by binding to ABCB1. In addition, circ_0002060 sponged miR-198 to upregulate ABCB1 expression. Conclusions: circ_0002060 promoted DOX resistance and OS progression by regulating the miR-198/ABCB1 axis, suggesting that circ_0002060 might be a promising biomarker for OS therapy. [ABSTRACT FROM AUTHOR]

Objective The aim of this study was to optimize the currently used direct spectrophotometric serum prolidase enzyme activity (SPEA) assay method and compare its diagnostic accuracy with current precipitation and direct spectrophotometric assay methods, AST-to-ALT ratio, age platelet index, AST-to-platelet ratio index, cirrhosis discriminate score, Doha score, FIB-4, FibroQ, fibrosis index, Goteborg University Cirrhosis Index , King's score, and Pohl score for distinguishing Ishak F0 from F1–F3 in patients with chronic hepatitis B (CHB) infection. Methods Liver biopsy results from 112 patients were included in this study.  Results The SPEA values were 529 (292–794) U/L, 671 (486–927) U/L, and 1077 (867–1399) U/L with the precipitation, current, and optimized direct spectrophotometric assay methods, respectively. According to multivariate logistic regression analysis optimized direct spectrophotometric SPEA was the only statistically significant parameter to predict the early stages of liver fibrosis.  Conclusions Optimized direct spectrophotometric SPEA assay method could be used to distinguish early stages of liver fibrosis in patients with CHB infection instead of the currently used spectrophotometric SPEA assay methods and other evaluated liver fibrosis indexes. [ABSTRACT FROM AUTHOR]

Simple Summary: Triple-negative breast cancer (TNBC) has a high recurrence rate and metastasis rate, and the prognosis for patients is poor. Identifying novel treatment avenues for TNBC is crucial for the treatment and prognosis of TNBC patients. Deubiquitinase deficiency is associated with the occurrence and progression of several diseases, including cancer. As a member of deubiquitinase, ubiquitin-specific peptidase 53 (USP53) is essential for the development of tumors such as hepatocellular carcinoma and renal clear cell carcinoma. However, the precise function of USP53 in TNBC remains unknown. Our study has identified USP53 as a molecular target that mediates the progression of TNBC. This not only deepens our understanding of the tumorigenesis and development of TNBC, but also provides new potential therapeutic targets for TNBC patients. Breast cancer (BC) ranks in the top five malignant tumors in terms of morbidity and mortality rates. Among BC subtypes, TNBC has a high recurrence rate and metastasis rate and the worst prognosis. However, the exact mechanism by which TNBC develops is unclear. Here, we show that the deubiquitinase USP53 contributes to tumor growth and metastasis in TNBC. USP53 is overexpressed in TNBC, and this phenotype is linked to a poor prognosis. Functionally, USP53 promotes TNBC cell proliferation, migration, invasion, and epithelial–mesenchymal transition (EMT). More importantly, USP53 decreases the chemosensitivity of BC cells by enhancing v-crk sarcoma virus CT10 oncogene homologue (avian)-like (CRKL) expression. Mechanistically, USP53 directly binds CRKL to stabilize and deubiquitinate it, thereby preventing CRKL degradation. Overall, we discovered that USP53 deubiquitinates CRKL, encourages tumor development and metastasis, and reduces chemosensitivity in TNBC. These findings imply that USP53 might represent a new therapeutic target for the treatment of TNBC. [ABSTRACT FROM AUTHOR]

Simple Summary: Malignant pleural mesothelioma belongs to the aggressive tumor of pleura commonly recognized at the advanced, chemoresistant stage. Thymidylate synthase (TYMS) is a critical therapeutic target because it determines the susceptibility to folate-related drug treatment. We have previously reported that a natural extract isolated from defatted evening primrose seeds (EPE) reduces cell proliferation and the invasiveness of MPM cells. Therefore, we aimed to expound the mechanism of EPE in downregulating TYMS expression. Our bioinformatic analysis of an MPM patient genes data set confirmed the correlation of overexpressed TYMS with epithelial-to-mesenchymal transition biomarkers. Mechanistic studies of the culture cell have provided evidence for the direct effect of EPE compounds on the transcriptional activity of tyms promoter to its transcription factor, specificity protein 1. EPE mixture is a complex intervention related to the occupancy of SP1 motifs, blocking TYMS expression and triggering invasiveness of MPM cells. Purpose: To determine the mechanism of EPE in downregulating TYMS in MPM cancer. Methods: The TYMS mRNA expression with epithelial-to-mesenchymal transition biomarkers and nuclear factor SP1 was assessed using the GEO database in a data set of MPM patients (GSE51024). Invasive MPM cell lines were in vitro models for the investigation of TYMS expression after EPE treatment. The tyms promoter SP1 binding sequences were determined using Genomatix v 3.4 software Electrophoretic mobility shift and dual-luciferase reporter assays revealed specific SP1 motifs in the interaction of EPE and reference compounds. Chromatin immunoprecipitation and Re-ChIP were used for the co-occupancy study. Results: In MPM patients, a positive correlation of overexpressed TYMS with mesenchymal TWIST1, FN1 and N-cadherin was observed. EPE and its major components, gallic and ellagic acid (GA and EA, respectively), downregulated TYMS in invasive MPM cells by interacting with particular SP1 motifs on the tyms promoter. The luciferase constructs confirmed the occupation of two SP1 regulatory regions critical for the promotion of TYMS expression. Both EPE and reference standards influenced SP1 translocation into the nucleus. Conclusion: EPE components reduced TYMS expression by occupation of SP1 motifs on the tyms promoter and reversed the EMT phenotype of invasive MPM cells. Further in-depth analysis of the molecular docking of polyphenol compounds with SP1 regulatory motifs is required. [ABSTRACT FROM AUTHOR]

Simple Summary: The progression of PDAC is also regulated at the transcription/transduction stage. RNA metabolism is manipulated by different RNA-binding proteins (RBPs), and insulin-like growth factor 2 mRNA-binding proteins (IMPs), such as IMP1, have been associated with a poor prognosis in PDAC patients; however, little is known about its contribution to PDAC carcinogenesis. Public data suggest that PDAC patients with higher IMP1 expression showed poor overall survival. IMP1 silencing leads to reduced cell growth in PDAC cells and three-dimensional spheroids. Abrogation of IMP1 in PDAC cells showed lower levels of CDC25A, increased phosphorylation of the cyclin-dependent kinase (CDK)2, and accumulation of cells in the G1 phase. Immunoprecipitation revealed that IMP1 binds CDC25A mRNA, thereby controlling cell cycle progression. Ultimately, we proved that suppression of IMP1 blocked in vivo growth of Panc-1 transferred into immunodeficient mice. Our results indicate that IMP1 drives the PDCA cell cycle and represents a novel strategy for overcoming PDCA cell proliferation. A number of data indicate that the sources of different kinds of PDAC may be discovered at the transcription/transduction stage. RNA metabolism is manipulated at various steps by different RNA-binding proteins (RBPs), and the deregulation or irregular activity of RBPs is known to contribute to tumor promotion and progression. The insulin-like growth factor 2 mRNA-binding protein family (IMPs), and IMP1 in particular, has been linked with a poor prognosis in PDAC patients; however, little is known about its contribution in PDAC carcinogenesis. In this study, we investigated the function of IMP1 in PDAC. To evaluate IMP1 expression and correlation with PDAC prognosis, we utilized several public databases. Using a specific siRNA IMP1, we analyzed cell death and cell cycle progression in PDAC cell lines and 3D spheroids. The role of IMP1 was also evaluated in vivo in a Panc-1-derived tumor xenograft murine model. Public data suggest that PDAC patients with higher expression of IMP1 showed poor overall and progression-free survival. IMP1 silencing leads to reduced cell growth in PDAC cells and three-dimensional spheroids. Abrogation of IMP1 in PDAC cells showed lower levels of CDC25A, increased phosphorylation of the cyclin-dependent kinase (CDK)2, and accumulation of PDAC cells in the G1 phase. Immunoprecipitation experiments revealed that IMP1 binds CDC25A mRNA, thus controlling cell-cycle progression. Ultimately, we proved that suppression of IMP1 blocked in vivo growth of Panc-1 transferred into immunodeficient mice. Our results indicate that IMP1 drives the PDCA cell cycle and represents a novel strategy for overcoming PDCA cell proliferation. [ABSTRACT FROM AUTHOR]

Simple Summary: Pancreatic ductal adenocarcinoma (PDAC) is a tough cancer. Instead of stopping certain genes, we used their power to change tumors. Surprisingly, some proteins that usually help the cancer grow slowed it down when they were outside the cells. We found that one protein, called PSCA, which is usually bad for patients, could help if we placed it extracellularly. We conducted experiments to see how the cancer cells reacted, and we also looked at how a conditioned medium from the blood could affect the cancer. We found that PSCA could make the cancer cells weaker and less able to spread; it works together with other medicines that are used to fight cancer. This discovery could lead to new ways to treat cancer by using the same protein that was causing the issue. Overall, our study shows that some things that have a negative impact could be used to help fight this tough cancer. Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with low survival rates. We explored an innovative therapeutic approach by leveraging prognostic oncogenic markers. Instead of inhibiting these marker genes, we harnessed their tumor-modifying potential in the extracellular domain. Surprisingly, many of the proteins highly expressed in PDAC, which is linked to poor survival, exhibited tumor-suppressing qualities in the extracellular environment. For instance, prostate stem cell antigens (PSCA), associated with reduced survival, acted as tumor suppressors when introduced extracellularly. We performed in vitro assays to assess the proliferation and migration and evaluated the tumor-modifying capacity of extracellular factors from peripheral blood mononuclear cells (PBMCs) in PDAC tissues. Molecular docking analysis, immunoprecipitation, Western blotting, and RNA interference were employed to study the regulatory mechanism. Extracellular PSCA recombinant protein notably curtailed the viability, motility, and transwell invasion of PDAC cells. Its anti-PDAC effects were partially mediated by Mesothelin (MSLN), another highly expressed tumor-associated antigen in PDAC. The anti-tumor effects of extracellular PSCA complemented those of chemotherapeutic agents like Irinotecan, 5-Fluorouracil, and Oxaliplatin. PSCA expression increased in a conditioned medium derived from PBMCs and T lymphocytes. This study unveils the paradoxical anti-PDAC potential of PSCA, hinting at the dual roles of oncoproteins like PSCA in PDAC suppression. [ABSTRACT FROM AUTHOR]

Background: The aim of this study was to explore the function and mechanism of circular RNA (circRNA) matrix metallopeptidase 1 (circMMP1) in the progression of esophageal squamous cell carcinoma (ESCC). Methods: CircMMP1 expression was detected by quantitative real‐time PCR (qRT‐PCR), and its relationship with the prognosis of ESCC patients was evaluated by Kaplan–Meier analysis. Cells were transfected using corresponding plasmids, and the cell proliferation activity, migration and invasion capabilities in vitro were assessed. The protein level in tissues and cells was analyzed using western blotting. RNA pulldown, dual‐luciferase reporter assay and RNA immunoprecipitation assay were performed in ESCC cells to detect the interaction between circMMP1 and miR‐671‐5p, or the correlation between miR‐671‐5p and ANO1. Xenograft tumor experiment was carried out to uncover the function of circMMP1 in vivo. Results: The high level of circMMP1 in tumor tissues was associated with poor prognoses of ESCC patients. Knockdown of circMMP1 suppressed ESCC cell proliferation, migration and invasion in vitro. MiR‐671‐5p was the target of circMMP1 and mediated the inhibition effect of circMMP1 on ESCC cells. CircMMP1 targeted miR‐671‐5p to regulate ANO1 expression, which was downstream of miR‐671‐5p. Overexpression of ANO1 weakened tumor‐repressive function of circMMP1 knockdown in ESCC cells. Moreover, silencing of circMMP1 impeded ESCC tumor growth in vivo. Conclusion: Our study provided novel evidence that circMMP1 accelerated ESCC progression by acting as a miR‐671‐5p sponge to enhance ANO1 expression. [ABSTRACT FROM AUTHOR]

Objectives: Stem cells of the apical papilla (SCAPs) provide promising candidates for dental pulp regeneration. Despite great advances in the transcriptional controls of the SCAPs fate, little is known about the regulation of SCAP differentiation. Materials and Methods: Short hairpin RNAs and full‐length RNA were used to deplete or overexpress lysine demethylase 4D (KDM4D) gene expression. Western blotting, real‐time RT‐PCR, alizarin red staining, and scratch migration assays were used to study the role of KDM4D and the ribosomal protein encoded by RPS5 in SCAPs. RNA microarray, chromatin Immunoprecipitation (ChIP), and co‐immunoprecipitation (Co‐IP) assays were performed to explore the underlying molecular mechanisms. Results: KDM4D enhanced the osteo/dentinogenic differentiation, migration, and chemotaxis of SCAPs. The microarray results revealed that 88 mRNAs were differentially expressed in KDM4D‐overexpressed SCAPs. ChIP results showed knock‐down of KDM4D increased the level of H3K9me2 and H3K9me3 in CNR1 promoter region. There were 37 possible binding partners of KDM4D. KDM4D was found to combine with RPS5, which also promoted the osteo/dentinogenic differentiation, migration, and chemotaxis of SCAPs. Conclusions: KDM4D promoted the osteo/dentinogenic differentiation and migration potential of SCAPs in combination with RPS5, which provides a therapeutic clue for improving SCAPs‐based dental tissue regeneration. [ABSTRACT FROM AUTHOR]

Simple Summary: The protein TAZ plays an important role in the Hippo signaling pathway that regulates important biological processes such as cell proliferation, controlled cell death, cancer stem cell traits, tumorigenesis, and resistance to therapies. Specifically, TAZ functions as a transcription coactivator that influences the transcription of various genes. Despite the importance of TAZ, genome-wide occupancy sites for TAZ remain poorly defined. Here, a tetracycline (tet)-inducible gene expression system was used to turn on TAZ activation in mammary epithelial cells, and we characterized genome-wide binding sites at different TAZ activation time points. We found that most TAZ binds to distant enhancer and non-coding regions of genes at earlier TAZ activation time points and binding shifts to nearby promoter regions of genes at later TAZ activation time points. We also found that TAZ activation results in chromatin architecture alterations. These results could lead to the identification of new therapeutic targets for breast cancer. The transcriptional co-activator with PDZ binding motif (TAZ) is a key effector of the Hippo signaling pathway. We and others previously reported that high expression levels of TAZ are positively associated with decreased survival rates and shorter times to relapse in basal-like breast cancer (BLBC) patients. The oncogenic activity of TAZ involves the regulation of diverse signal transduction pathways that direct processes such as cell proliferation, migration, and resistance to apoptosis, albeit through poorly characterized gene expression programs. Here, using a tet-inducible system in mammary epithelial MCF10A cells, we have characterized the TAZ-regulated transcription program using RNA sequencing in a temporal and spatial manner. We further identified global TAZ binding sites at different TAZ activation time points by chromatin immunoprecipitation (ChIP) sequencing analysis. We found that the vast majority of TAZ was rapidly localized in enhancer regions at the early TAZ activation time point and then gradually spread to promoter regions. TAZ bound to enhancer regions following a switch in potential TEAD and FOSL2 transcription factor motifs. Furthermore, the ATAC sequencing analysis indicated that TAZ activation led to chromatin structural alterations. Together, our results have revealed the landscape of genome-wide TAZ binding sites and may lead to improvements in the current understanding of how TAZ regulates the gene expression program that contributes to the development of breast cancer. [ABSTRACT FROM AUTHOR]

Background: mTORC1 (mechanistic target of rapamycin complex 1) is associated with lymphoma progression. Oncogenic RRAGC (Rag guanosine triphosphatase C) mutations identified in patients with follicular lymphoma facilitate the interaction between Raptor (regulatory protein associated with mTOR) and Rag GTPase. It promotes the activation of mTORC1 and accelerates lymphomagenesis. Cardamonin inhibits mTORC1 by decreasing the protein level of Raptor. In the present study, we investigated the inhibitory effect and possible mechanism of action of cardamonin in RRAGC-mutant lymphoma. This could provide a precise targeted therapy for lymphoma with RRAGC mutations. Methods: Cell viability was measured using a cell counting kit-8 (CCK-8) assay. Protein expression and phosphorylation levels were determined using western blotting. The interactions of mTOR and Raptor with RagC were determined by co-immunoprecipitation. Cells overexpressing RagC wild-type (RagCWT) and RagC Thr90Asn (RagCT90N) were generated by lentiviral infection. Raptor knockdown was performed by lentivirus-mediated shRNA transduction. The in vivo anti-tumour effect of cardamonin was assessed in a xenograft model. Results: Cardamonin disrupted mTOR complex interactions by decreasing Raptor protein levels. RagCT90N overexpression via lentiviral infection increased cell proliferation and mTORC1 activation. The viability and tumour growth rate of RagCT90N-mutant cells were more sensitive to cardamonin treatment than those of normal and RagCWT cells. Cardamonin also exhibited a stronger inhibitory effect on the phosphorylation of mTOR and p70 S6 kinase 1 in RagCT90N-mutant cells. Raptor knockdown abolishes the inhibitory effects of cardamonin on mTOR. An in vivo xenograft model demonstrated that the RagCT90N-mutant showed significantly higher sensitivity to cardamonin treatment. Conclusions: Cardamonin exerts selective therapeutic effects on RagCT90N-mutant cells. Cardamonin can serve as a drug for individualised therapy for follicular lymphoma with RRAGC mutations. [ABSTRACT FROM AUTHOR]

Introduction: Proteolytic processing of amyloid protein precursor by b-site secretase enzyme (BACE1) is dependent on the cellular lipid composition and is affected by endomembrane trafficking in dementia and Alzheimer's disease (AD). Stearoyl-CoA desaturase 1 (SCD1) is responsible for the synthesis of fatty acid monounsaturation (MUFAs), whose accumulation is strongly associated with cognitive dysfunction. Methods: In this study, we analyzed the relationship between BACE1 and SCD1 in vivo and in vitro neurodegenerative models and their association in familial AD (FAD), sporadic AD (SAD), and cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) using microscopy, biochemical, and mass SPECT approach. Results: Our findings showed that BACE1 and SCD1 immunoreactivities were increased and colocalized in astrocytes of the hippocampus in a rat model of global cerebral ischemia (2-VO). A synergistic effect of double BACE1/SCD1 silencing on the recovery of motor and cognitive functions was obtained. This neuroprotective regulation involved the segregation of phospholipids (PLs) associated with polyunsaturated fatty acids in the hippocampus, cerebrospinal fluid, and serum. The double silencing in the sham and ischemic groups was stronger in the serum, inducing an inverse ratio between total phosphatydilcholine (PC) and lysophosphatidylcholine (LPC), representedmainly by the reduction of PC 38:4 and PC 36:4 and an increase in LPC 16:0 and LPC 18:0. Furthermore, PC 38:4 and PC:36:4 levels augmented in pathological conditions in in vitro AD models. BACE1 and SCD1 increases were confirmed in the hippocampus of FAD, SAD, and CADASIL. Conclusion: Therefore, the findings suggest a novel convergence of BACE-1 and SCD1 in neurodegeneration, related to pro-inflammatory phospholipids. [ABSTRACT FROM AUTHOR]

Objective: To investigate the mechanism of RNA-binding protein hnRNP A1 in mouse hippocampal neurons (HT22) on glycolysis. Methods: RIP and CLIP-qPCR were performed by HT22 in vitro to observe the mechanism of hnRNP A1 regulating the expression of key proteins in glycolysis. The RNA binding domain of hnRNP A1 protein in HT22 was inhibited by VPC-80051, and the effect of hnRNP A1 on glycolysis of HT22 was observed. Lentivirus overexpression of hnRNP A1 was used to observe the effect of overexpression of hnRNP A1 on glycolysis of Aβ25-35-injured HT22. The expression of hnRNP A1 in brain tissues of wild-type mice and triple-transgenic (APP/PS1/Tau) AD mice at different ages was studied by Western blot assay. Results: The results of RIP experiment showed that hnRNP A1 and HK1 mRNA were significantly bound. The results of CLIP-qPCR showed that hnRNP A1 directly bound to the 2605-2821 region of HK1 mRNA. hnRNP A1 inhibitor can down-regulate the expression of HK1 mRNA and HK1 protein in HT22 cells. Overexpression of hnRNP A1 can significantly reduce the toxic effect of Aβ25-35 on neurons via the hnRNP A1/HK1/pyruvate pathway. In addition, inhibition of hnRNP A1 binding to amyloid precursor protein (APP) RNA was found to increase A25-35 expression, while Aβ25-35 also down-regulated hnRNP A1 expression by enhancing phosphorylation of p38 MAPK in HT22. They interact to form bidirectional regulation, further down-regulating the expression of hnRNP A1, and ultimately aggravating glycolytic dysfunction. Protein immunoblotting showed that hnRNP A1 decreased with age in mouse brain tissue, and the decrease was greater in AD mice, suggesting that the decrease of hnRNP A1 may be a predisposed factor in the pathogenesis of AD. [ABSTRACT FROM AUTHOR]

Objective The presence of FMF cases without MEFV (MEFV innate immunity regulator, pyrin) pathogenic variants led us to search for other genes' involvement in the disease development. Here, we describe the presence of genetic heterogeneity in a three-generation family with an FMF/mevalonate kinase deficiency (MKD)-overlapping phenotype without MEFV/MVK (mevalonate kinase) pathogenic variants. Method Targeted sequencing revealed a rare, fully penetrant variant in PSTPIP1 (p.Arg228Cys, rs781341816). Computational stability analyses of PSTPIP1 protein were performed. PSTPIP1-pyrin protein interaction was examined by immunoprecipitation and immunoblotting in peripheral blood mononuclear cells (PBMCs) of patients and healthy controls. PBMCs were cultured, and inflammation was induced by LPS+ATP treatment, followed by protein level measurements of caspase-1, IL1ß, pyrin and PSTPIP1 in cell lysates and mature caspase-1 and mature IL1ß in supernatants. Results The conserved, rare (GnomAD, 0.000028) PSTPIP1 p.Arg228Cys variant, previously reported in ClinVar as a variant with uncertain significance, showed complete penetrance in the family presenting an autosomal dominant pattern. Computational analyses showed a potentially destabilizing effect of the variant on PSTPIP1 protein. Accordingly, PSTPIP1-pyrin interaction was increased in patients harboring the variant, which resulted in elevated levels of mature caspase-1 and IL1ß in the inflammation-induced patient samples. Conclusions Unlike previously described cases with pyogenic arthritis, pyoderma gangrenosum, and acne (PAPA)-associated PSTPIP1 variants, our patients with the p.Arg228Cys variant presented with an FMF/MKD-overlapping phenotype. As additional data on the genetic heterogeneity in the variable clinical spectrum of autoinflammatory syndromes, we suggest that the p.Arg228Cys variant in PSTPIP1 is related to inflammation responses through strong PSTPIP1 -pyrin interaction and pyrin inflammasome activation. [ABSTRACT FROM AUTHOR]

Ischemia/reperfusion (I/R) is a pathological process that causes severe damage. Propofol is known to alleviate I/R-related injury; however, the exact function and underlying mechanisms are not fully understood. Using an oxygen glucose deprivation/re-oxygenation (OGD/R) method, an in vitro I/R injury model was induced. The cell viability and the level of Fe2+, glutathione synthetase (GSH), and malondialdehyde (MDA) were evaluated using kits. Luciferase reporter gene assay, chromatin immunoprecipitation, and RNA immunoprecipitation (RIP) were used to verify the interaction between molecules. The m6A level of BECN1 mRNA was determined through methylated RIP. Propofol-treated OGD/R models showed reduced levels of Fe2+ and MDA, while the cell viability and the level of GSH increased. Propofol inhibited ferroptosis by down-regulating HIF-1α in OGD/R-treated HT22 cells. HIF-1α is bound to the promoter region of YTHDF1 to promote its transcription, and YTHDF1 promoted ferroptosis by stabilizing the mRNA of BECN1. The suppressive effect of propofol on OGD/R-induced ferroptosis was reversed by the overexpression of YTHDF1. Our study revealed that the HIF-1α/YTHDF1/BECN1 axis in OGD/R-treated HT22 cells promotes ferroptosis, and administration of propofol can inhibit this axis to avoid cell death. This study provides a novel insight for the neuroprotective function of propofol. [ABSTRACT FROM AUTHOR]

BACKGROUND: Several epidemiological investigations demonstrated that maternal arsenic (As) exposure elevated risk of fetal growth restriction (FGR), but the mechanism remains unclear. OBJECTIVES: This study aimed to investigate the effects of gestational As exposure on placental and fetal development and its underlying mechanism. METHODS: Dams were exposed to 0.15, 1.5, and 15 mg/L NaAsO2 throughout pregnancy via drinking water. Sizes of fetuses and placentas, placental histopathology, and glycogen content were measured. Placental RNA sequencing was conducted. Human trophoblasts were exposed to NaAsO2 (2 μM) to establish an in vitro model of As exposure. The mRNA stability and protein level of genes identified through RNA sequencing were measured. 푁6-Methyladenosine (m6A) modification was detected by methylated RNA immunoprecipitation–quantitative real-time polymerase chain reason (qPCR). The binding ability of insulin-like growth factor 2 binding protein 2 to the gene of interest was detected by RNA-binding protein immunoprecipitation-qPCR. Intracellular S-adenosylmethionine (SAM) and methyltransferase activity were determined by liquid chromatography– tandem mass spectrometry (LC-MS/MS) and colorimetry, respectively. In vitro As+3 methyltransferase (As3MT) knockdown or SAM supplementation and in vivo folic acid (FA) supplementation were used to evaluate the protective effect. A case–control study verified the findings. RESULTS: Sizes of fetuses (exposed to 1.5 and 15 mg/L NaAsO2) and placentas (exposed to 15 mg/L NaAsO2) were lower in As-exposed mice. More glycogen+ trophoblasts accumulated and the expression of markers of interstitial invasion was lower in the 15 mg/L NaAsO2-exposed mouse group in comparison with control. Placental RNA sequencing identified cysteine-rich angiogenic inducer 61 (Cyr61) as a candidate gene of interest. Mechanistically, mice and cells exposed to As had lower protein expression of CYR61, and this was attributed to a lower incidence of Cyr61 m6A. Furthermore, cells exposed to As had lower methyltransferase activity, suggesting that this could be the mechanism by which Cyr61 m6A was affected. Depletion of intracellular SAM, a cofactor for m6A methyltransferase catalytic domain, partially contributed to As-induced methyltransferase activity reduction. Either As3MT knockdown or SAM supplementation attenuated As-induced Cyr61 m6A down-regulation. In mice, FA supplementation rescued As-induced defective trophoblastic invasion and FGR. In humans, a negative correlation between maternal urinary As and plasma CYR61 was observed in infants who were small for gestational age. DISCUSSION: Using in vitro and in vivo models, we found that intracellular SAM depletion–mediated Cyr61 m6A down-regulation partially contributed to As-induced defective trophoblastic invasion and FGR. [ABSTRACT FROM AUTHOR]