Regulatory Factors Research Articles

Roles of G proteins and their GTPase-activating proteins in platelets.

Platelets are small anucleate blood cells supporting vascular function. They circulate in a quiescent state monitoring the vasculature for injuries. Platelets adhere to injury sites and can be rapidly activated to secrete granules and to form platelet/platelet aggregates. These responses are controlled by signalling networks that include G proteins and their regulatory guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). Recent proteomics studies have revealed the complete spectrum of G proteins, GEFs, and GAPs present in platelets. Some of these proteins are specific for platelets and very few have been characterised in detail. GEFs and GAPs play a major role in setting local levels of active GTP-bound G proteins in response to activating and inhibitory signals encountered by platelets. Thus, GEFs and GAPs are highly regulated themselves and appear to integrate G protein regulation with other cellular processes. This review focuses on GAPs of small G proteins of the Arf, Rab, Ras, and Rho families, as well as of heterotrimeric G proteins found in platelets.

A novel non-canonical function for IRF6 in the recycling of E-cadherin.

Interferon Regulatory Factor 6 (IRF6) is a transcription factor essential for keratinocyte cell-cell adhesions. Previously, we found that recycling of E-cadherin was defective in the absence of IRF6, yet total E-cadherin levels were not altered, suggesting a previously unknown, non-transcriptional function for IRF6. IRF6 protein contains a DNA binding domain (DBD) and a protein binding domain (PBD). The transcriptional function of IRF6 depends on its DBD and PBD, however, whether the PBD is necessary for the interaction with cytoplasmic proteins has yet to be demonstrated. Here, we show that an intact PBD is required for recruitment of cell-cell adhesion proteins at the plasma membrane, including the recycling of E-cadherin. Colocalizations and co-immunoprecipitations reveal that IRF6 forms a complex in recycling endosomes with Rab11, Myosin Vb and E-cadherin, and that the PBD is required for this interaction. These data indicate that IRF6 is a novel effector of the endosomal recycling of E-cadherin and demonstrate a non-transcriptional function for IRF6 in regulating cell-cell adhesions.

The novel regulator HdrR controls the transcription of the heterodisulfide reductase operon hdrBCA in Methanosarcina barkeri.

Methanogenic archaea play a key role in the global carbon cycle because these microorganisms remineralize organic compounds in various anaerobic environments. The microorganism Methanosarcina barkeri is a metabolically versatile methanogen, which can utilize acetate, methanol, and H2/CO2 to synthesize methane. However, the regulatory mechanisms underlying methanogenesis for different substrates remain unknown. In this study, RNA-seq analysis was used to investigate M. barkeri growth and gene transcription under different substrate regimes. According to the results, M. barkeri showed the best growth under methanol, followed by H2/CO2 and acetate, and these findings corresponded well with the observed variations in genes transcription abundance for different substrates. In addition, we identified a novel regulator, MSBRM_RS03855 (designated as HdrR), which specifically activates the transcription of the heterodisulfide reductase hdrBCA operon in M. barkeri. HdrR was able to bind to the hdrBCA operon promoter to regulate transcription. Furthermore, the structural model analyses revealed a helix-turn-helix domain, which is likely involved in DNA binding. Taken together, HdrR serves as a model to reveal how certain regulatory factors control the expression of key enzymes in the methanogenic pathway.IMPORTANCEThe microorganism Methanosarcina barkeri has a pivotal role in the global carbon cycle and contributes to global temperature homeostasis. The consequences of biological methanogenesis are far-reaching, including impacts on atmospheric methane and CO2 concentrations, agriculture, energy production, waste treatment, and human health. As such, reducing methane emissions is crucial to meeting set climate goals. The methanogenic activity of certain microorganisms can be drastically reduced by inhibiting the transcription of the hdrBCA operon, which encodes heterodisulfide reductases. Here, we provide novel insight into the mechanisms regulating hdrBCA operon transcription in the model methanogen M. barkeri. The results clarified that HdrR serves as a regulator of heterodisulfide reductase hdrBCA operon transcription during methanogenesis, which expands our understanding of the unique regulatory mechanisms that govern methanogenesis. The findings presented in this study can further our understanding of how genetic regulation can effectively reduce the methane emissions caused by methanogens.

DNA methylase 1 influences temperature responses and development in the invasive pest Tuta absoluta.

DNA methylase 1 (Dnmt1) is an important regulatory factor associated with biochemical signals required for insect development. It responds to changes in the environment and triggers phenotypic plasticity. Meanwhile, Tuta absolutaMeyrick (Lepidoptera: Gelechiidae)-a destructive invasive pest-can rapidly invade and adapt to different habitats; however, the role of Dnmt1 in this organism has not been elucidated. Accordingly, this study investigates the mechanism(s) underlying the rapid adaptation of Tuta absoluta to temperature stress. Potential regulatory genes were screened via RNAi (RNA interference), and the DNA methylase in Tuta absoluta was cloned by RACE (Rapid amplification of cDNA ends). TaDnmt1 was identified as a potential regulatory gene via bioinformatics; its expression was evaluated in response to temperature stress and during different development stages using real-time polymerase chain reaction. Results revealed that TaDnmt1 participates in hot/cold tolerance, temperature preference and larval development. The full-length cDNA sequence of TaDnmt1 is 3765 bp and encodes a 1254 kDa protein with typical Dnmt1 node-conserved structural features and six conserved DNA-binding active motifs. Moreover, TaDnmt1 expression is significantly altered by temperature stress treatments and within different development stages. Hence, TaDnmt1 likely contributes to temperature responses and organismal development. Furthermore, after treating with double-stranded RNA and exposing Tuta absoluta to 35°C heat shock or -12°C cold shock for 1 h, the survival rate significantly decreases; the preferred temperature is 2°C lower than that of the control group. In addition, the epidermal segments become enlarged and irregularly folded while the surface dries up. This results in a significant increase in larval mortality (57%) and a decrease in pupation (49.3%) and eclosion (50.9%) rates. Hence, TaDnmt1 contributes to temperature stress responses and temperature perception, as well as organismal growth and development, via DNA methylation regulation. These findings suggest that the rapid geographic expansion of T absoluta has been closely associated with TaDnmt1-mediated temperature tolerance. This study advances the research on 'thermos Dnmt' and provides a potential target for RNAi-driven regulation of Tuta absoluta.

Trends in hospital price transparency after implementation of the CMS Final Rule.

To assess trends in hospital price disclosures after the Centers for Medicare & Medicaid Services (CMS) Final Rule went into effect. The Turquoise Health Price Transparency Dataset was used to identify all US hospitals that publicly displayed pricing from 2021 to 2023. Price-disclosing versus nondisclosing hospitals were compared using Pearson's Chi-squared and Wilcoxon rank sum tests. Bayesian structural time-series modeling was used to determine if enforcement of increased penalties for nondisclosure was associated with a change in the trend of hospital disclosures. Not applicable. As of January 2023, 5162 of 6692 (77.1%) US hospitals disclosed pricing of their services, with the majority (2794 of 5162 [54.1%]) reporting their pricing within the first 6 months of the final rule going into effect in January 2021. An increase in hospital disclosures was observed after penalties for nondisclosure were enforced in January 2022 (relative effect size 20%, p = 0.002). Compared with nondisclosing hospitals, disclosing hospitals had higher annual revenue, bed number, and were more likely to be have nonprofit ownership, academic affiliation, provide emergency services, and be in highly concentrated markets (p < 0.001). Hospital pricing disclosures are continuously in flux and influenced by regulatory and market factors.

JrPHL8-JrWRKY4-JrSTH2L module regulates resistance to Colletotrichum gloeosporioides in walnut

Abstract Walnut anthracnose (Colletotrichum gloeosporioides) reduces walnut yield and quality and seriously threatens the healthy development of the walnut industry. WRKY transcription factors (TFs) are crucial regulatory factors involved in plant-pathogen interactions. Our previous transcriptome analysis results indicate that JrWRKY4 responds to infection by C. gloeosporioides, but its specific regulatory network and disease resistance mechanism are still unclear. Herein, the characteristics of JrWRKY4 as a transcription activator located in the nucleus were first identified. Gain-of-function and loss-of-function analyses showed that JrWRKY4 could enhance walnut resistance against C. gloeosporioides. A series of molecular experiments showed that JrWRKY4 directly interacted with the promoter region of JrSTH2L and positively regulated its expression. In addition, JrWRKY4 interacted with JrVQ4 to form the protein complex, which inhibited JrWRKY4 for the activation of JrSTH2L. Notably, a MYB TF JrPHL8 interacting with the JrWRKY4 promoter was also been identified, which directly bound to the MBS element in the promoter of JrWRKY4 and induced its activity. Our study elucidated a novel mechanism of the JrPHL8-JrWRKY4-JrSTH2L in regulating walnut resistance to anthracnose. This mechanism improves our understanding of the molecular mechanism of WRKY TF mediated resistance to anthracnose in walnut, which provides new insights for molecular breeding of disease-resistant walnuts in the future.

MiR-106a targets ATG7 to inhibit autophagy and angiogenesis after myocardial infarction.

Myocardial infarction (MI) is an acute condition in which the heart muscle dies due to the lack of blood supply. Previous research has suggested that autophagy and angiogenesis play vital roles in the prevention of heart failure after MI, and miR-106a is considered to be an important regulatory factor in MI. But the specific mechanism remains unknown. In this study, using cultured venous endothelial cells and a rat model of MI, we aimed to identify the potential target genes of miR-106a and discover the mechanisms of inhibiting autophagy and angiogenesis. We first explored the biological functions of miR-106a on autophagy and angiogenesis on endothelial cells. Then we identified ATG7, which was the downstream target gene of miR-106a. The expression of miR-106a and ATG7 was investigated in the rat model of MI. We found that miR-106a inhibits the proliferation, cell cycle, autophagy and angiogenesis, but promoted the apoptosis of vein endothelial cells. Moreover, ATG7 was identified as the target of miR-106a, and ATG7 rescued the inhibition of autophagy and angiogenesis by miR-106a. The expression of miR-106a in the rat model of MI was decreased but the expression of ATG7 was increased in the infarction areas. Our results indicate that miR-106a may inhibit autophagy and angiogenesis by targeting ATG7. This mechanism may be a potential therapeutic treatment for MI.

Single-cell RNA-binding protein pattern-mediated molecular subtypes depict the hallmarks of the tumor microenvironment in bladder urothelial carcinoma

Abstract Objectives Bladder carcinoma (BC) is a common malignancy of the urinary tract. As a new hallmark of cancer for drug therapy, RNA-binding proteins (RBPs) are key regulatory factors in alternative splicing events. This work is to uncover the relationship between BC and RBP in order to find drug targets in BC. Methods In this work, data from single-cell RNA-seq GSE1355337, PRJNA662018, and the TCGA-Bladder urothelial carcinoma (BLCA) cohorts are integrated to identify their relationships. A scoring system is constructed according to RBPs gene expression and patients’ survival. A network is constructed to analyze the alternative splicing events and RBP genes. Results A scoring system identified 321 RBPs significantly associated with the prognosis of patients. Subsequent typing of these RBP genes in two single-cell datasets demonstrated that most of the RBP genes had variable copy numbers. Three RBP clusters were identified. Using RBP genes as a signature in BC epithelial cells allows for differentiation between different grades of BC samples. The novel RBP genes-based subtype system reflects BC clinical staging. Notably, CellChat analysis revealed that the RBP genes-associated cell subtypes of T cells had extensive interactions with epithelial cells. Further analysis showed that the ligand-receptor pair MIF-CXCR4 mediated the communication between RBP-associated subtypes of BC epithelial cells and T cells. Conclusions Taken together, RBP genes are associated with BC progress and offer new indicators for precision medicine in BC.

Studying the Pathogenic Effect of New Nucleotide Changes in the Promoter Region of the TERT Gene in Patients with Glioblastoma Brain Tumor

Introduction: Brain tumors are considered to be one of the most dangerous types of cancer, despite decades of research and treatment efforts. The activation of the telomerase enzyme is a critical factor in the immortality of these cells. Mutations in the promoter region of the TERT gene, particularly in the promoter hot spots, can influence the binding of activating transcription factors and inhibit the binding of negative regulatory factors of the TERT gene, ultimately regulating the gene's expression. This study aimed to identify and investigate the mutations of the TERT gene promoter region in glioblastoma, a malignant brain tumor, using bioinformatics. Methods: A study was conducted using the case-control method where 35 patients with glioblastoma multiform brain tumor were examined along with 40 people who were examined as control samples. In this research, the Touchdown PCR technique and direct DNA sequencing were used to detect mutations in the promoter region of the TERT gene in patients with glioblastoma. Furthermore, bioinformatic analyses were conducted to investigate the pathogenic effect of nucleotide changes in this gene region. Results: In the core region of the TERT gene promoter, three-point mutations were identified (c.*146C&gt;T، c.*144C&gt;G and c.*143G&gt;C). Two of these mutations were novel and have been observed for the first time in glioblastoma multiform. The remaining mutation was located in the promoter core's hot spot of the gene. This mutation was known as c.*146C&gt;T. Conclusion: In this research, the harmful impact of TERT promoter region mutations as a biomarker for glioblastoma was examined. These findings suggest that mutations in regulatory regions, as well as coding sequences, could be significant contributory factors in the process of carcinogenesis.

Genome-Wide Transcriptional Roles of KSHV Viral Interferon Regulatory Factors in Oral Epithelial Cells

Genome-Wide Transcriptional Roles of KSHV Viral Interferon Regulatory Factors in Oral Epithelial Cells

Application of Computational Law and Artificial Intelligence Methods for Sharia Compliance Analysis of E-Waste Management Systems Based on Blockchain

This paper aims to 1) develop a methodology using computational law and blockchain technology to ensure Sharia compliance in e-waste management systems; 2) propose a conceptual model for a Sharia blockchain platform enabling compliance monitoring and verification; and 3) formulate recommendations to facilitate real-world implementation. The study employs a conceptual approach using inductive analysis of Sharia principles, deductive reasoning, and systems modeling to formulate design requirements and components for a blockchain platform that can enable transparent, tamper-proof monitoring and control of e-waste handling in accordance with Islamic law. Firstly, analysis reveals five key criteria for Sharia compliance in e-waste management and how blockchain, smart contracts, and AI can address these. A comprehensive compliance assurance methodology leveraging these technologies is proposed. Secondly, a conceptual model is delineated for a multipurpose Sharia blockchain platform encompassing consensus, cryptography, smart contract rule engines, and AI analytics. Thirdly, recommendations are synthesized covering technological, regulatory, economic, infrastructure, and social factors needed to enable real-world implementation. This pioneering research bridges Sharia law, sustainability, and emerging technologies to offer culturally attuned e-waste management solutions. The methodology, conceptual models, and practical framework devised inform development of next-generation systems for values-aligned compliance assurance, with potential impact extending beyond the realm of e-waste contexts.

Nur77-IRF1 axis inhibits esophageal squamous cell carcinoma growth and improves anti-PD-1 treatment efficacy

The nuclear receptor Nur77 plays paradoxical roles in numerous cancers. However, whether Nur77 inhibits esophageal squamous cell carcinoma (ESCC) growth and affects immunological responses against ESCC has not been determined. The functional role of Nur77 in ESCC was investigated in this study using human ESCC cell lines, quantitative real-time polymerase chain reaction (PCR), cell proliferation and colony formation assays, flow cytometry analysis, western blotting and animal models. The target gene controlled by Nur77 was verified using dual-luciferase reporter assays, chromatin immunoprecipitation analysis and functional rescue experiments. To examine the clinical importance of Nur77, 72 human primary ESCC tissues were subjected to immunohistochemistry. Taken together, these findings showed that, both in vitro and in vivo, Nur77 dramatically reduced ESCC cell growth and triggered apoptosis. Nur77 directly interacts with the interferon regulatory factor 1 (IRF1) promoter to inhibit its activity in ESCC. Pharmacological induction of Nur77 using cytosporone B (CsnB) inhibited ESCC cell proliferation and promoted apoptosis both in vitro and in vivo. Furthermore, CsnB increased CD8+ T-cell infiltration and cytotoxicity to inhibit the formation of ESCC tumors in an immunocompetent mouse model. In ESCC tissues, Nur77 expression was downregulated, and IRF1 expression was increased; moreover, their expression levels were negatively related. IRF1 and Nur77 were strongly correlated with overall survival. These findings suggested that Nur77 targets and regulates the IRF1/PD-L1 axis to serve as a tumor suppressor in ESCC.Graphical abstract of the regulatory mechanism of Nur77 overexpression downregulates IRF1 in the inhibition of ESCC progression and enhance anti-PD-1 therapy efficacy.

Inflammation in HIV and Its Impact on Atherosclerotic Cardiovascular Disease.

People living with HIV have a 1.5- to 2-fold increased risk of developing cardiovascular disease. Despite treatment with highly effective antiretroviral therapy, people living with HIV have chronic inflammation that makes them susceptible to multiple comorbidities. Several factors, including the HIV reservoir, coinfections, clonal hematopoiesis of indeterminate potential (CHIP), microbial translocation, and antiretroviral therapy, may contribute to the chronic state of inflammation. Within the innate immune system, macrophages harbor latent HIV and are among the prominent immune cells present in atheroma during the progression of atherosclerosis. They secrete inflammatory cytokines such as IL (interleukin)-6 and tumor necrosis-α that stimulate the expression of adhesion molecules on the endothelium. This leads to the recruitment of other immune cells, including cluster of differentiation (CD)8+ and CD4+ T cells, also present in early and late atheroma. As such, cells of the innate and adaptive immune systems contribute to both systemic inflammation and vascular inflammation. On a molecular level, HIV-1 primes the NLRP3 (NLR family pyrin domain containing 3) inflammasome, leading to an increased expression of IL-1β, which is important for cardiovascular outcomes. Moreover, activation of TLRs (toll-like receptors) by HIV, gut microbes, and substance abuse further activates the NLRP3 inflammasome pathway. Finally, HIV proteins such as Nef (negative regulatory factor) can inhibit cholesterol efflux in monocytes and macrophages through direct action on the cholesterol transporter ABCA1 (ATP-binding cassette transporter A1), which promotes the formation of foam cells and the progression of atherosclerotic plaque. Here, we summarize the stages of atherosclerosis in the context of HIV, highlighting the effects of HIV, coinfections, and antiretroviral therapy on cells of the innate and adaptive immune system and describe current and future interventions to reduce residual inflammation and improve cardiovascular outcomes among people living with HIV.

The E3 ubiquitin-protein ligase UHRF1 promotes adipogenesis and limits fibrosis by suppressing GPNMB-mediated TGF-β signaling

The E3 ubiquitin-ligase UHRF1 is an epigenetic regulator coordinating DNA methylation and histone modifications. However, little is known about how it regulates adipogenesis or metabolism. In this study, we discovered that UHRF1 is a key regulatory factor for adipogenesis, and we identified the altered molecular pathways that UHRF1 targets. Using CRISPR/Cas9-based knockout strategies, we discovered the whole transcriptomic changes upon UHRF1 deletion. Bioinformatics analyses revealed that key adipogenesis regulators such PPAR-γ and C/EBP-α were suppressed, whereas TGF-β signaling and fibrosis markers were upregulated in UHRF1-depleted differentiating adipocytes. Furthermore, UHRF1-depleted cells showed upregulated expression and secretion of TGF-β1, as well as the glycoprotein GPNMB. Treating differentiating preadipocytes with recombinant GPNMB led to an increase in TGF-β protein and secretion levels, which was accompanied by an increase in secretion of fibrosis markers such as MMP13 and a reduction in adipogenic conversion potential. Conversely, UHRF1 overexpression studies in human cells demonstrated downregulated levels of GPNMB and TGF-β, and enhanced adipogenic potential. In conclusion, our data show that UHRF1 positively regulates 3T3-L1 adipogenesis and limits fibrosis by suppressing GPNMB and TGF-β signaling cascade, highlighting the potential relevance of UHRF1 and its targets to the clinical management of obesity and linked metabolic disorders.

Glycolysis and beyond in glucose metabolism: exploring pulmonary fibrosis at the metabolic crossroads

At present, pulmonary fibrosis (PF) is a prevalent and irreversible lung disease with limited treatment options, and idiopathic pulmonary fibrosis (IPF) is one of its most common forms. Recent research has highlighted PF as a metabolic-related disease, including dysregulated iron, mitochondria, lipid, and glucose homeostasis. Systematic reports on the regulatory roles of glucose metabolism in PF are rare. This study explores the intricate relationships and signaling pathways between glucose metabolic processes and PF, delving into how key factors involved in glucose metabolism regulate PF progression, and the interplay between them. Specifically, we examined various enzymes, such as hexokinase (HK), 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3 (PFKFB3), pyruvate kinase (PK), and lactate dehydrogenase (LDH), illustrating their regulatory roles in PF. It highlights the significance of lactate, alongside the role of pyruvate dehydrogenase kinase (PDK) and glucose transporters (GLUTs) in modulating pulmonary fibrosis and glucose metabolism. Additionally, critical regulatory factors such as transforming growth factor-beta (TGF-β), interleukin-1 beta (IL-1β), and hypoxia-inducible factor 1 subunit alpha (HIF-1α) were discussed, demonstrating their impact on both PF and glucose metabolic pathways. It underscores the pivotal role of AMP-activated protein kinase (AMPK) in this interplay, drawing connections between diabetes mellitus, insulin, insulin-like growth factors, and peroxisome proliferator-activated receptor gamma (PPARγ) with PF. This study emphasizes the role of key enzymes, regulators, and glucose transporters in fibrogenesis, suggesting the potential of targeting glucose metabolism for the clinical diagnosis and treatment of PF, and proposing new promising avenues for future research and therapeutic development.

Analysis of 14–3-3 Gene Family and Functional Characterization of DlGRF8/11 modulating flowering time in Ma bamboo (Dendrocalamus latiflorus Munro)

14–3–3 proteins, also known as general regulatory factors (GRFs), are common regulatory proteins that play a critical role in growth, development, and environmental responses in plants. However, there is limited understanding of the members and functions of 14–3–3 gene family in ma bamboo. In this study, 25 Dl14–3–3 genes were identified and designated as DlGRF1-DlGRF25, respectively. Bioinformatics analysis of DlGRF genes provided insights into the conservation and diversity during evolution in ma bamboo. It’s verified that the expression patterns of DlGRFs were discrepant in different flowering tissues. To further explore the floral transition, Y2H, BiFC, and LCI assays revealed that DlGRF8 interacted with DlFT1/DlTFL1 in the cytoplasm and with DlFT2/DlFD1 in the nucleus. Interestingly, DlGRF11 exclusively interacted with DlFD1 in the nucleus. Overexpressing experiments in rice indicated that DlGRF8 promoted flowering, while DlGRF11 delayed flowering. Moreover, they exhibited diverse expression patterns in response to drought and ABA treatments. These results provide preliminary insights into the role of DlGRF genes in floral transition, growth, and development.

#2416 Bicc1 and PKD: linking RNA metabolism to kidney disease

Abstract Background and Aims Mutations in PKD1 or PKD2 lead to autosomal dominant polycystic kidney disease (PKD) by allowing urine flow-sensing primary cilia to activate tissue growth signals. However, the responsible cilia-associated factors and a strategy to block them and cure PKD are elusive. Cilia also specify organ laterality. In this process, flow-stimulated cilia activate the cytosolic protein Bicaudal-C (Bicc1) to curb the translation and/or the stability of specific target mRNAs. To identify candidate regulatory factors, we investigated if Bicc1 ribonucleoprotein particle (RNP) formation involves known proteins linked to cilia and cystic kidney diseases. Method To address this question, we here used AlphaFold structure modeling of protein complexes, combined with in vitro reconstitution assays and live imaging of Bicc1 RNP assembly. To begin to validate these interactions, we tested by genetic approaches if specific interacting factors also regulate Bicc1 targets in cells and mice. Results We found that multivalent interactions with structured domains of the ankyrin repeat and SAM domain containing (ANKS) 3 and their allosteric regulation by the related ANKS6 protein inhibit or restore Bicc1 access to target mRNAs, respectively. Concomitantly, ANKS3 alone increased the solubility and fluidity of Bicc1, whereas co-recruitment of ANKS6 had the opposite effect, liberating Bicc1 to self-polymerize and to induce phase-transitioning of target mRNAs in gel-like meshworks. Conclusion Perturbations in Bicc1-ANKS3-ANKS6 RNP dynamics and altered translation of Bicc1 target mRNAs emerge as likely culprits regulating cyst formation downstream of ciliary defects. ANKS6 can be sequestered in cilia by inversin, and mutations in these proteins provoke kidney cysts in nephronophthisis patients. Therefore, besides systematically profiling Bicc1 RNPs, future studies should test if ANKS6 mediates their dynamic regulation by flow-stimulated cilia.

Exosomes derived from miR-146a-overexpressing fibroblast-like synoviocytes in cartilage degradation and macrophage M1 polarization: a novel protective agent for osteoarthritis?

IntroductionPathological changes in the articular cartilage (AC) and synovium are major manifestations of osteoarthritis (OA) and are strongly associated with pain and functional limitations. Exosome-derived microRNAs (miRNAs) are crucial regulatory factors in intercellular communication and can influence the progression of OA by participating in the degradation of chondrocytes and the phenotypic transformation in the polarization of synovial macrophages. However, the specific relationships and pathways of action of exosomal miRNAs in the pathological progression of OA in both cartilage and synovium remain unclear.MethodsThis study evaluates the effects of fibroblast-like synoviocyte (FLS)-derived exosomes (FLS-Exos), influenced by miR-146a, on AC degradation and synovial macrophage polarization. We investigated the targeted relationship between miR-146a and TRAF6, both in vivo and in vitro, along with the involvement of the NF-κB signaling pathway.ResultsThe expression of miR-146a in the synovial exosomes of OA rats was significantly higher than in healthy rats. In vitro, the upregulation of miR-146a reduced chondrocyte apoptosis, whereas its downregulation had the opposite effect. In vivo, exosomes derived from miR-146a-overexpressing FLSs (miR-146a-FLS-Exos) reduced AC injury and chondrocyte apoptosis in OA. Furthermore, synovial proliferation was reduced, and the polarization of synovial macrophages shifted from M1 to M2. Mechanistically, the expression of TRAF6 was inhibited by targeting miR-146a, thereby modulating the Toll-like receptor 4/TRAF6/NF-κB pathway in the innate immune response.DiscussionThese findings suggest that miR-146a, mediated through FLS-Exos, may alleviate OA progression by modulating cartilage degradation and macrophage polarization, implicating the NF-κB pathway in the innate immune response. These insights highlight the therapeutic potential of miR-146a as a protective agent in OA, underscoring the importance of exosomal miRNAs in the pathogenesis and potential treatment of the disease.

#921 Interferon regulatory factor 4 (IRF4)-specific deletion on dendritic cells protects against kidney inflammation and injury in IRI-induced AKI/AKD

Abstract Background and Aims Ischemia-reperfusion injury (IRI) is a frequent clinical cause of acute kidney injury and disease (AKI/AKD) which can lead to acute tubular necrosis and irreversible kidney function loss. Mononuclear phagocytes, including dendritic cells (DCs), Monocytes, and macrophages, play important roles in various phases of injury and repair processes. Previous research has indicated that conventional dendritic cells (cDCs) are subdivided into cDC1s and cDC2s, and these two subsets could have opposing functions. We also demonstrated that cDC1s have a protective role as deficiency of cDC1s results in aggravated inflammation and injury in the IRI-induced AKI/AKD model in our previous study. Based on these researches, we aim to investigate the function of cDC2s in the IRI-induced AKI/AKD model. Method We used interferon regulatory factor 4 (IRF4)-deficient mice (IRF4fl/flClec9acre) to specifically delete IRF4 on DC lineage cells as our experimental group, and used wide-type mice as our control group. After unilateral IRI (UIRI) surgery, we measured the GFR and plasma creatinine of the mice on day 1 (D1) and day 5 (D5), then collected the injured kidney issue after sacrificing mice. We compared the differences between IRF4fl/flClec9acre mice and wild-type mice in terms of DC subpopulation changes, neutrophil accumulation, T cell differentiation, intensity of inflammatory responses, and tissue repair capabilities in damaged kidney tissue, by using techniques such as immunohistochemistry, immunofluorescence, flow cytometry, qRT-PCR, etc. Results ([KOMean-WTMean] ± SEM): IRF4fl/flClec9acre mice showed a lower percentage of cDC2s without affecting cDC1s in the kidney compared with wide-type mice (D1: −0.04467 ± 0.01356, P = 0.0301; D5: −1.108 ± 0.2248, P = 0.0004). During post-ischemic AKI/AKD, IRF4 deficiency in cDCs had several beneficial effects compared to wild-type mice, including reducing renal tubular cell injury (PAS score D1: −0.9375 ± 0.1523, P &amp;lt; 0.0001; PAS score D5: −1.119 ± 0.1445, P &amp;lt; 0.0001), decreasing loss of kidney function (GFR D1: 8.983 ± 4.091, P = 0.1041; GFR D5: 14.9 ± 5.01, P = 0.0172), and promoting kidney function recovery. This was associated with increased expression of anti-inflammatory cytokines (IL-4 D1: 4.486 ± 1.363, P = 0.0079; IL-4 D5: 3.403 ± 1.447, P = 0.0366; IL-10 D1: 2.103 ± 0.7342, P = 0.0299; IL-10 D5: 2.654 ± 0.8859, P = 0.0166), decreased expression of pro-inflammatory cytokines(TNF-α D1: −1.418 ± 0.5467, P = 0.0433; TNF-α D5: −2.434 ± 1.919, P = 0.0074; IL-23a D1: −1.320 ± 0.3432, P = 0.0061; IL-23a D5: −16.12 ± 3.587, P = 0.0004) and acute tubular cell death, reduced neutrophil chemoattractant expression (CXCL-2) (Fig. 6) and recruitment of neutrophils (−6.107 ± 1.664, P = 0.0214), and differentiation of Th2 (−0.1867 ± 0.03333, P = 0.005) and Th17 cells (−0.1767 ± 0.01333, P = 0.0002). Conclusion In conclusion, our data demonstrate a critical role of IRF4 in the kidney cDC2 subset and highlight that deficiency of cDC2s can protect against kidney inflammation, reduce kidney injury, and promote kidney recovery in the IRI-induced AKI/AKD model. This suggests that cDC2s have an immunoregulatory role in AKI/AKD and could serve as a potential therapeutic target for kidney inflammation.

Pathologic TDP-43 downregulates myelin gene expression in the monkey brain.

Growing evidence indicates that non-neuronal oligodendrocyte plays an important role in Amyotrophic lateral sclerosis (ALS) and other neurodegenerative diseases. In patient's brain, the impaired myelin structure is a pathological feature with the observation of TDP-43 in cytoplasm of oligodendrocyte. However, the mechanism underlying the gain of function by TDP-43 in oligodendrocytes, which are vital for the axonal integrity, remains unclear. Recently, we found that the primate-specific cleavage of truncated TDP-43 fragments occurred in cytoplasm of monkey neural cells. This finding opened up the avenue to investigate the myelin integrity affected by pathogenic TDP-43 in oligodendrocytes. In current study, we demonstrated that the truncated TDP-35 in oligodendrocytes specifically, could lead to the dysfunctional demyelination in corpus callosum of monkey. As a consequence of the interaction of myelin regulatory factor with the accumulated TDP-35 in cytoplasm, the downstream myelin-associated genes expression was downregulated at the transcriptional level. Our study aims to investigate the potential effect on myelin structure injury, affected by the truncated TDP-43 in oligodendrocyte, which provided the additional clues on the gain of function during the progressive pathogenesis and symptoms in TDP-43 related diseases.