RESUMO
BACKGROUND: N-acetyltransferase 10 (NAT10) plays a crucial role in the occurrence and development of various tumors. However, the current regulatory mechanism of NAT10 in tumors is limited to its presence in tumor cells. Here, we aimed to reveal the role of NAT10 in intrahepatic cholangiocarcinoma (ICC) and investigate its effect on macrophage polarization in the tumor microenvironment (TME). METHODS: The correlation between NAT10 and ICC clinicopathology was analyzed using tissue microarray (TMA), while the effect of NAT10 on ICC proliferation was verified in vitro and in vivo. Additionally, the downstream target of NAT10, C-C motif chemokine ligand 2 (CCL2), was identified by Oxford Nanopore Technologies full-length transcriptome sequencing, RNA immunoprecipitation-quantitative polymerase chain reaction, and coimmunoprecipitation experiments. It was confirmed by co-culture that ICC cells could polarize macrophages towards M2 type through the influence of NAT10 on CCL2 protein expression level. Through RNA-sequencing, molecular docking, and surface plasmon resonance (SPR) assays, it was confirmed that berberine (BBR) can specifically bind CCL2 to inhibit ICC development. RESULTS: High expression level of NAT10 was associated with poor clinicopathological manifestations of ICC. In vitro, the knockdown of NAT10 inhibited the proliferative activity of ICC cells and tumor growth in vivo, while its overexpression promoted ICC proliferation. Mechanically, by binding to CCL2 messenger RNA, NAT10 increased CCL2 protein expression level in ICC and their extracellular matrix, thereby promoting the proliferation of ICC cells and M2-type polarization of macrophages. BBR can target CCL2, inhibit ICC proliferation, and reduce M2-type polarization of macrophages. CONCLUSIONS: NAT10 promotes ICC proliferation and M2-type polarization of macrophages by up-regulating CCL2, whereas BBR inhibits ICC proliferation and M2-type polarization of macrophages by inhibiting CCL2.
Assuntos
Proliferação de Células , Quimiocina CCL2 , Colangiocarcinoma , Macrófagos , Quimiocina CCL2/metabolismo , Colangiocarcinoma/patologia , Colangiocarcinoma/genética , Colangiocarcinoma/metabolismo , Macrófagos/metabolismo , Humanos , Animais , Linhagem Celular Tumoral , Neoplasias dos Ductos Biliares/patologia , Neoplasias dos Ductos Biliares/genética , Neoplasias dos Ductos Biliares/metabolismo , Masculino , Microambiente Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Polaridade Celular/efeitos dos fármacos , Camundongos Nus , Camundongos , Pessoa de Meia-Idade , Ligação ProteicaRESUMO
Spinal microglial polarization plays a crucial role in the pathological processes of neuropathic pain following peripheral nerve injury. Accumulating evidence suggests that milk fat globule epidermal growth factor-8 (MFG-E8) exhibits anti-inflammatory effect and regulates microglial polarization through the integrin ß3 receptor. However, the impact of MFG-E8 on microglial polarization in the context of neuropathic pain has not yet been investigated. In this study, we evaluated the effect of MFG-E8 on pain hypersensitivity and spinal microglial polarization following spared nerve injury (SNI) of the sciatic nerve in mice. We determined the molecular mechanisms underlying the effects of MFG-E8 on pain hypersensitivity and spinal microglial polarization using pain behavior assessment, western blot (WB) analysis, immunofluorescence (IF) staining, quantitative polymerase chain reaction (qPCR), enzyme-linked immunosorbent assay (ELISA), and small interfering RNA (siRNA) transfection. Our findings indicate that SNI significantly increased the levels of MFG-E8 and integrin ß3 expressed in microglia within the spinal cord of mice. Additionally, we observed that intrathecal injection of recombinant human MFG-E8 (rhMFG-E8) alleviated SNI induced-mechanical allodynia and thermal hyperalgesia. Furthermore, the results suggested that rhMFG-E8 facilitated M2 microglial polarization and ameliorated neuroinflammation via integrin ß3/SOCS3/STAT3 pathway in the spinal cord of mice with SNI. Importantly, these effects were negated by integrin ß3 siRNA, or SOCS3 siRNA. These results demonstrate that MFG-E8 ameliorates peripheral nerve injury induced-mechanical allodynia and thermal hyperalgesia by driving M2 microglial polarization and mitigating neuroinflammation mediated by integrin ß3/SOCS3/STAT3 pathway in the spinal cord of mice. MFG-E8 may serve as a promising target for the treatment of neuropathic pain.
Assuntos
Antígenos de Superfície , Integrina beta3 , Microglia , Proteínas do Leite , Neuralgia , Fator de Transcrição STAT3 , Transdução de Sinais , Proteína 3 Supressora da Sinalização de Citocinas , Animais , Camundongos , Microglia/metabolismo , Proteína 3 Supressora da Sinalização de Citocinas/metabolismo , Antígenos de Superfície/metabolismo , Neuralgia/metabolismo , Integrina beta3/metabolismo , Integrina beta3/biossíntese , Masculino , Fator de Transcrição STAT3/metabolismo , Proteínas do Leite/biossíntese , Transdução de Sinais/fisiologia , Camundongos Endogâmicos C57BL , Doenças Neuroinflamatórias/metabolismo , Traumatismos dos Nervos Periféricos/metabolismo , Traumatismos dos Nervos Periféricos/complicações , Polaridade Celular/fisiologia , Polaridade Celular/efeitos dos fármacosRESUMO
OBJECTIVE: To investigate the molecular mechanism by which DNA damage induced apoptosis suppressor (DDIAS) regulates STAT3/CCL2 to enhance macrophage polarization to M1 type in Kawasaki disease (KD). METHODS: A KD vascular model was established by culturing human coronary artery endothelial cells (HCAECs) in vitro. Small interfering RNA of DDIAS (si-DDIAS) was transfected into the KD cell model. The human macrophage cell line THP-1 was induced into M1 macrophages using phorbol myristate acetate (PMA) and lipopolysaccharide (LPS) and co-cultured with the endothelial cells using the HCAECs medium. Western blot analysis was utilized to assess cellular DDIAS, p-STAT3, STAT3, and CCL2 protein expression. MTT was utilized to detect cell proliferation. ELISA was utilized to assess the expression levels of TNF-α, IL-4, IL-6, IL-8 and CCL2 in cell supernatants. Flow cytometry was utilized to examine cell apoptosis and the expression of M1 macrophage surface marker CD86. RESULTS: The expression level of DDIAS was elevated in the KD group compared to the Control group. Serum inhibition of HCAEC proliferation in the KD group was concentration-dependent and pro-inflammatory cytokines were substantially elevated, while the anti-inflammatory cytokines were substantially reduced (P<0.05). Compared to the si-NC group, cell proliferation was considerably enhanced; pro-inflammatory cytokines were substantially reduced; anti-inflammatory cytokines were substantially elevated, and the expression of p-STAT3 and CCL2 was lowered in the si-DDIAS group (P<0.05). The percentage of M1 macrophages was substantially elevated in the THP-1+LPS group compared to the THP-1 group (P<0.05). Compared to the THP-1+LPS+si-NC group, macrophage CCL2 expression was decreased in the THP-1+LPS+si-DDIAS group; the percentage of M1 macrophages was substantially lowered (P<0.05); and the levels of pro-inflammatory cytokines and CCL2 in the cell supernatant were substantially reduced. Incubation of macrophages with STAT3 agonist reversed these changes, which were exacerbated by the addition of neutralizing antibody CCL2. CONCLUSIONS: Downregulation of DDIAS inhibits macrophage polarization toward the M1 type through inhibition of the STAT3/CCL2 signaling pathway and can ameliorate vascular injury and inflammation in KD coronary arteries.
Assuntos
Quimiocina CCL2 , Macrófagos , Síndrome de Linfonodos Mucocutâneos , Fator de Transcrição STAT3 , Humanos , Fator de Transcrição STAT3/metabolismo , Síndrome de Linfonodos Mucocutâneos/metabolismo , Síndrome de Linfonodos Mucocutâneos/patologia , Quimiocina CCL2/metabolismo , Macrófagos/metabolismo , Apoptose , Proliferação de Células , Células Endoteliais/metabolismo , Vasos Coronários/patologia , Vasos Coronários/metabolismo , Citocinas/metabolismo , Células THP-1 , Polaridade Celular/efeitos dos fármacosRESUMO
Pharyngeal endoderm cells undergo convergence and extension (C&E), which is essential for endoderm pouch formation and craniofacial development. Our previous work implicates Gα13/RhoA-mediated signaling in regulating this process, but the underlying mechanisms remain unclear. Here, we have used endoderm-specific transgenic and Gα13 mutant zebrafish to demonstrate that Gα13 plays a crucial role in pharyngeal endoderm C&E by regulating RhoA activation and E-cadherin expression. We showed that during C&E, endodermal cells gradually establish stable cell-cell contacts, acquire apical-basal polarity and undergo actomyosin-driven apical constriction, which are processes that require Gα13. Additionally, we found that Gα13-deficient embryos exhibit reduced E-cadherin expression, partially contributing to endoderm C&E defects. Notably, interfering with RhoA function disrupts spatial actomyosin activation without affecting E-cadherin expression. Collectively, our findings identify crucial cellular processes for pharyngeal endoderm C&E and reveal that Gα13 controls this through two independent pathways - modulating RhoA activation and regulating E-cadherin expression - thus unveiling intricate mechanisms governing pharyngeal endoderm morphogenesis.
Assuntos
Caderinas , Endoderma , Subunidades alfa G12-G13 de Proteínas de Ligação ao GTP , Regulação da Expressão Gênica no Desenvolvimento , Faringe , Proteínas de Peixe-Zebra , Peixe-Zebra , Proteína rhoA de Ligação ao GTP , Animais , Endoderma/metabolismo , Endoderma/embriologia , Endoderma/citologia , Caderinas/metabolismo , Caderinas/genética , Peixe-Zebra/embriologia , Peixe-Zebra/metabolismo , Peixe-Zebra/genética , Proteína rhoA de Ligação ao GTP/metabolismo , Proteína rhoA de Ligação ao GTP/genética , Proteínas de Peixe-Zebra/metabolismo , Proteínas de Peixe-Zebra/genética , Subunidades alfa G12-G13 de Proteínas de Ligação ao GTP/metabolismo , Subunidades alfa G12-G13 de Proteínas de Ligação ao GTP/genética , Faringe/embriologia , Faringe/metabolismo , Actomiosina/metabolismo , Transdução de Sinais , Morfogênese/genética , Polaridade Celular , Animais Geneticamente Modificados , Embrião não Mamífero/metabolismoRESUMO
Polarized exocytosis induced by local Cdc42 GTPase activity results in membrane flows that deplete low-mobility membrane-associated proteins. A reaction-diffusion particle model comprising Cdc42 positive feedback activation, hydrolysis by GTPase-activating proteins (GAPs), and flow-induced displacement by exo/endocytosis shows that flow-induced depletion of low mobility GAPs promotes polarization. We modified Cdc42 mobility in Schizosaccharomyces pombe by replacing its prenylation site with 1, 2 or 3 repeats of the Rit C-terminal membrane-binding domain (ritC), yielding alleles with progressively lower mobility and increased flow-coupling. While Cdc42-1ritC cells are viable and polarized, Cdc42-2ritC polarize poorly and Cdc42-3ritC are inviable, in agreement with model's predictions. Deletion of Cdc42 GAPs restores viability to Cdc42-3ritC cells, verifying the model's prediction that GAP deletion increases Cdc42 activity at the expense of polarization. Our work demonstrates how membrane flows are an integral part of Cdc42-driven pattern formation and require Cdc42-GTP to turn over faster than the surface on which it forms.
Assuntos
Membrana Celular , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces , Proteína cdc42 de Ligação ao GTP , Schizosaccharomyces/metabolismo , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Proteína cdc42 de Ligação ao GTP/metabolismo , Proteína cdc42 de Ligação ao GTP/genética , Membrana Celular/metabolismo , Polaridade Celular , Proteínas Ativadoras de GTPase/metabolismo , Proteínas Ativadoras de GTPase/genética , Forma Celular , Exocitose/fisiologia , EndocitoseRESUMO
BACKGROUND: C-X-C receptor 4(CXCR4) is widely considered to be a highly conserved G protein-coupled receptor, widely involved in the pathophysiological processes in the human body, including fibrosis. However, its role in regulating macrophage-related inflammation in the fibrotic process of prostatitis has not been confirmed. Here, we aim to describe the role of CXCR4 in modulating macrophage M1 polarization through glycolysis in the development of prostatitis fibrosis. METHODS: Use inducible experimental chronic prostatitis as a model of prostatic fibrosis. Reduce CXCR4 expression in immortalized bone marrow-derived macrophages using lentivirus. In the fibrotic mouse model, use adenovirus carrying CXCR4 agonists to detect the silencing of CXCR4 and assess the in vivo effects. RESULTS: In this study, we demonstrated that reducing CXCR4 expression during LPS treatment of macrophages can alleviate M1 polarization. Silencing CXCR4 can inhibit glycolytic metabolism, enhance mitochondrial function, and promote macrophage transition from M1 to M2. Additionally, in vivo functional experiments using AAV carrying CXCR4 showed that blocking CXCR4 in experimental autoimmune prostatitis (EAP) can alleviate inflammation and experimental prostate fibrosis development. Mechanistically, CXCR4, a chemokine receptor, when silenced, weakens the PI3K/AKT/mTOR pathway as its downstream signal, reducing c-MYC expression. PFKFB3, a key enzyme involved in glucose metabolism, is a target gene of c-MYC, thus impacting macrophage polarization and glycolytic metabolism processes.
Assuntos
Fibrose , Glicólise , Macrófagos , Próstata , Receptores CXCR4 , Masculino , Animais , Receptores CXCR4/metabolismo , Receptores CXCR4/genética , Macrófagos/metabolismo , Camundongos , Próstata/patologia , Próstata/metabolismo , Prostatite/patologia , Prostatite/metabolismo , Prostatite/genética , Transdução de Sinais , Camundongos Endogâmicos C57BL , Humanos , Serina-Treonina Quinases TOR/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Polaridade Celular , Fosfatidilinositol 3-Quinases/metabolismo , Fosfofrutoquinase-2/metabolismo , Fosfofrutoquinase-2/genéticaRESUMO
Primary osteoarthritis (OA) is a prevalent degenerative joint disease that mostly affects the knee joint. It is a condition that occurs around the world. Because of the aging population and the increase in obesity prevalence, the incidence of primary OA is increasing each year. Joint replacement can completely subside the pain and minimize movement disorders caused by advanced OA, while nonsteroidal drugs and injection of sodium hyaluronate into the joint cavity can only partially relieve the pain; hence, it is critical to search for new methods to treat OA. Increasing lines of evidence show that primary OA is a chronic inflammatory disorder, with synovial inflammation as the main characteristic. Macrophages, as one of the immune cells, can be polarized to produce M1 (proinflammatory) and M2 (anti-inflammatory) types during synovial inflammation in OA. Following polarization, macrophages do not come in direct contact with chondrocytes; however, they affect chondrocyte metabolism through paracrine production of a significant quantity of inflammatory cytokines, matrix metalloproteinases, and growth factors and thus participate in inducing joint pain, cartilage injury, angiogenesis, and osteophyte formation. The main pathways that influence the polarization of macrophages are the Toll-like receptor and NF-κB pathways. The study of how macrophage polarization affects OA disease progression has gradually become one of the approaches to prevent and treat OA. Experimental studies have found that the treatment of macrophage polarization in primary OA can effectively relieve synovial inflammation and reduce cartilage damage. The present article summarizes the influence of inflammatory factors secreted by macrophages after polarization on OA disease progression, the main signaling pathways that induce macrophage differentiation, and the role of different polarized types of macrophages in OA; thus, providing a reference for preventing and treating primary OA.
Assuntos
Progressão da Doença , Macrófagos , Osteoartrite , Humanos , Macrófagos/fisiologia , Osteoartrite/etiologia , Osteoartrite/patologia , Animais , Polaridade Celular/fisiologiaRESUMO
Ciliated epithelia are widespread in animals and play crucial roles in many developmental and physiological processes. Epithelia composed of multi-ciliated cells allow for directional fluid flow in the trachea, oviduct and brain cavities. Monociliated epithelia play crucial roles in vertebrate embryos, from the establishment of left-right asymmetry to the control of axis curvature via cerebrospinal flow motility in zebrafish. Cilia also have a central role in the motility and feeding of free-swimming larvae in a variety of marine organisms. These diverse functions rely on the coordinated orientation (rotational polarity) and asymmetric localization (translational polarity) of cilia and of their centriole-derived basal bodies across the epithelium, both being forms of planar cell polarity (PCP). Here, we review our current knowledge on the mechanisms of the translational polarity of basal bodies in vertebrate monociliated epithelia from the molecule to the whole organism. We highlight the importance of live imaging for understanding the dynamics of centriole polarization. We review the roles of core PCP pathways and of apicobasal polarity proteins, such as Par3, whose central function in this process has been recently uncovered. Finally, we emphasize the importance of the coordination between polarity proteins, the cytoskeleton and the basal body itself in this highly dynamic process.
Assuntos
Polaridade Celular , Centríolos , Cílios , Animais , Cílios/metabolismo , Cílios/fisiologia , Centríolos/metabolismo , Epitélio/metabolismo , Epitélio/fisiologia , Humanos , Células Epiteliais/metabolismo , Células Epiteliais/citologia , Corpos Basais/metabolismoRESUMO
Cell polarity - the asymmetric distribution of molecules and cell structures within the cell - is a feature that almost all cells possess. Even though the cytoskeleton and other intracellular organelles can have a direction and guide protein distribution, the plasma membrane is, in many cases, essential for the asymmetric localization of proteins because it helps to concentrate proteins and restrict their localization. Indeed, many proteins that exhibit asymmetric or polarized localization are either embedded in the PM or located close to it in the cellular cortex. Such proteins, which we refer to here as 'polar proteins', use various mechanisms of membrane targeting, including vesicle trafficking, direct phospholipid binding, or membrane anchoring mediated by post-translational modifications or binding to other proteins. These mechanisms are often shared with non-polar proteins, yet the unique combinations of several mechanisms or protein-specific factors assure the asymmetric distribution of polar proteins. Although there is a relatively detailed understanding of polar protein membrane targeting mechanisms in animal and yeast models, knowledge in plants is more fragmented and focused on a limited number of known polar proteins in different contexts. In this Review, we combine the current knowledge of membrane targeting mechanisms and factors for known plant transmembrane and cortical proteins and compare these with the mechanisms elucidated in non-plant systems. We classify the known factors as general or polarity specific, and we highlight areas where more knowledge is needed to construct an understanding of general polar targeting mechanisms in plants or to resolve controversies.
Assuntos
Membrana Celular , Polaridade Celular , Transporte Proteico , Membrana Celular/metabolismo , Animais , Proteínas de Plantas/metabolismo , Humanos , Proteínas de Membrana/metabolismoRESUMO
Adult mammalian lungs exhibit a fractal pattern, as each successive generation of airways is a fraction of the size of the parental branch. Achieving this structure likely requires precise control of airway length and diameter, as the embryonic airways initially lack the fractal scaling observed in the adult. In monolayers and tubes, directional growth can be regulated by the planar cell polarity (PCP) complex. Here, we characterized the roles of PCP complex components in airway initiation, elongation and widening during branching morphogenesis of the lung. Using tissue-specific knockout mice, we surprisingly found that branching morphogenesis proceeds independently of PCP complex function in the lung epithelium. Instead, we found a previously unreported Celsr1-independent role for the PCP complex components Vangl1 and Vangl2 in the pulmonary mesenchyme, where they are required for branch initiation, elongation and widening. Our data thus reveal an explicit function for Vangl1 and Vangl2 that is independent of the core PCP complex, suggesting a functional diversification of PCP complex components in vertebrate development. These data also reveal an essential role for the embryonic mesenchyme in generating the fractal structure of airways in the mature lung.
Assuntos
Polaridade Celular , Pulmão , Proteínas de Membrana , Mesoderma , Proteínas do Tecido Nervoso , Animais , Camundongos , Pulmão/embriologia , Pulmão/metabolismo , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Mesoderma/metabolismo , Mesoderma/embriologia , Camundongos Knockout , Morfogênese , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/genética , Receptores Acoplados a Proteínas GRESUMO
Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease characterized by chronic inflammation, lung tissue fibrotic changes and impaired lung function. Pulmonary fibrosis 's pathological process is thought to be influenced by macrophage-associated phenotypes. IPF treatment requires specific targets that target macrophage polarization. Cytokine-like 1(CYTL1) is a secreted protein with multiple biological functions first discovered in CD34+ haematopoietic cells. However, its possible effects on IPF progression remain unclear. This study investigated the role of CYTL1 in IPF progression in a bleomycin-induced lung injury and fibrosis model. In bleomycin-induced mice, CYTL1 is highly expressed. Moreover, CYTL1 ablation alleviates lung injury and fibrosis in vivo. Further, downregulating CYTL1 reduces macrophage M2 polarization. Mechanically, CYTL1 regulates transforming growth factor ß (TGF-ß)/connective tissue growth factor (CCN2) axis and inhibition of TGF-ß pathway alleviates bleomycin-induced lung injury and fibrosis. In conclusion, highly expressed CYTL1 inhibits macrophage M2 polarization by regulating TGF-ß/CCN2 expression, alleviating bleomycin-induced lung injury and fibrosis. CYTL1 could, therefore, serve as a promising IPF target.
Assuntos
Bleomicina , Fator de Crescimento do Tecido Conjuntivo , Regulação para Baixo , Macrófagos , Fibrose Pulmonar , Fator de Crescimento Transformador beta , Animais , Bleomicina/toxicidade , Camundongos , Regulação para Baixo/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismo , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/metabolismo , Fibrose Pulmonar/patologia , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Fator de Crescimento do Tecido Conjuntivo/genética , Camundongos Endogâmicos C57BL , Masculino , Polaridade Celular/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/metabolismo , Fibrose Pulmonar Idiopática/patologiaRESUMO
Bilateral communication between bones and muscles is essential for healing composite bone-muscle injuries from orthopedic surgeries and trauma. However, these injuries are often characterized by exaggerated inflammation, which can disrupt bone-muscle crosstalk, thereby seriously delaying the healing of either tissue. Existing approaches are largely effective at healing single tissues. However, simultaneous healing of multiple tissues remains challenging, with little research conducted to date. Here we introduce collagen patches that overcome this overlooked issue by harnessing the plasticity of macrophage phenotypes. Phosphatidylserine liposomes (PSLs) capable of shifting the macrophage phenotype from inflammatory M1 into anti-inflammatory/prohealing M2 were coated on collagen patches via a layer-by-layer method. Original collagen patches failed to improve tissue healing under inflammatory conditions coordinated by M1 macrophages. In contrast, PSL-coated collagen patches succeeded in accelerating bone and muscle healing by inducing a microenvironment dominated by M2 macrophages. In cell experiments, differentiation of preosteoblasts and myoblasts was completely inhibited by secretions of M1 macrophages but unaffected by those of M2 macrophages. RNA-seq analysis revealed that type I interferon and interleukin-6 signaling pathways were commonly upregulated in preosteoblasts and myoblasts upon stimulation with M1 macrophage secretions, thereby compromising their differentiation. This study demonstrates the benefit of PSL-mediated M1-to-M2 macrophage polarization for simultaneous bone and muscle healing, offering a potential strategy toward simultaneous regeneration of multiple tissues. STATEMENT OF SIGNIFICANCE: Existing approaches for tissue regeneration, which primarily utilize growth factors, have been largely effective at healing single tissues. However, simultaneous healing of multiple tissues remains challenging and has been little studied. Here we demonstrate that collagen patches releasing phosphatidylserine liposomes (PSLs) promote M1-to-M2 macrophage polarization and are effective for simultaneous healing of bone and muscle. Transcriptome analysis using next-generation sequencing reveals that differentiation of preosteoblasts and myoblasts is inhibited by the secretions of M1 macrophages but promoted by those of M2 macrophages, highlighting the importance of timely regulation of M1-to-M2 polarization in tissue regeneration. These findings provide new insight to tissue healing of multiple tissues.
Assuntos
Colágeno , Lipossomos , Macrófagos , Fosfatidilserinas , Lipossomos/química , Animais , Macrófagos/metabolismo , Macrófagos/efeitos dos fármacos , Fosfatidilserinas/metabolismo , Fosfatidilserinas/farmacologia , Camundongos , Colágeno/farmacologia , Colágeno/química , Cicatrização/efeitos dos fármacos , Células RAW 264.7 , Camundongos Endogâmicos C57BL , Diferenciação Celular/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Polaridade Celular/efeitos dos fármacos , Osteoblastos/metabolismo , Osteoblastos/efeitos dos fármacos , Osteoblastos/citologiaRESUMO
Developmental and epileptic encephalopathies (DEEs) feature altered brain development, developmental delay and seizures, with seizures exacerbating developmental delay. Here we identify a cohort with biallelic variants in DENND5A, encoding a membrane trafficking protein, and develop animal models with phenotypes like the human syndrome. We demonstrate that DENND5A interacts with Pals1/MUPP1, components of the Crumbs apical polarity complex required for symmetrical division of neural progenitor cells. Human induced pluripotent stem cells lacking DENND5A fail to undergo symmetric cell division with an inherent propensity to differentiate into neurons. These phenotypes result from misalignment of the mitotic spindle in apical neural progenitors. Cells lacking DENND5A orient away from the proliferative apical domain surrounding the ventricles, biasing daughter cells towards a more fate-committed state, ultimately shortening the period of neurogenesis. This study provides a mechanism for DENND5A-related DEE that may be generalizable to other developmental conditions and provides variant-specific clinical information for physicians and families.
Assuntos
Divisão Celular , Células-Tronco Pluripotentes Induzidas , Células-Tronco Neurais , Animais , Feminino , Humanos , Masculino , Camundongos , Polaridade Celular , Modelos Animais de Doenças , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Fatores de Troca do Nucleotídeo Guanina/genética , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Células-Tronco Neurais/metabolismo , Células-Tronco Neurais/citologia , Neurogênese/genéticaRESUMO
Cell polarity refers to the asymmetric distribution of proteins and other molecules along a specified axis within a cell. Polarity establishment is the first step in many cellular processes. For example, directed growth or migration requires the formation of a cell front and back. In many cases, polarity occurs in the absence of spatial cues. That is, the cell undergoes symmetry breaking. Understanding the molecular mechanisms that allow cells to break symmetry and polarize requires computational models that span multiple spatial and temporal scales. Here, we apply a multiscale modeling approach to examine the polarity circuit of yeast. In addition to symmetry breaking, experiments revealed two key features of the yeast polarity circuit: bistability and rapid dismantling of the polarity site following a loss of signal. We used modeling based on ordinary differential equations (ODEs) to investigate mechanisms that generate these behaviors. Our analysis revealed that a model involving positive and negative feedback acting on different time scales captured both features. We then extend our ODE model into a coarse-grained reaction-diffusion equation (RDE) model to capture the spatial profiles of polarity factors. After establishing that the coarse-grained RDE model qualitatively captures key features of the polarity circuit, we expand it to more accurately capture the biochemical reactions involved in the system. We convert the expanded model to a particle-based model that resolves individual molecules and captures fluctuations that arise from the stochastic nature of biochemical reactions. Our models assume that negative regulation results from negative feedback. However, experimental observations do not rule out the possibility that negative regulation occurs through an incoherent feedforward loop. Therefore, we conclude by using our RDE model to suggest how negative feedback might be distinguished from incoherent feedforward regulation.
Assuntos
Polaridade Celular , Modelos Biológicos , Saccharomyces cerevisiae , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Transdução de SinaisRESUMO
BACKGROUND: Spinal cord injury (SCI) is considered a central nervous system (CNS) disorder. Nuclear factor kappa B (NF-κB) regulates inflammatory responses in the CNS and is implicated in SCI pathogenesis. The mechanism(s) through which NF-κB contributes to the neuroinflammation observed during SCI however remains unclear. METHODS: SCI rat models were created using the weight drop method and separated into Sham, SCI and SCI+NF-κB inhibitor groups (n = 6 rats per-group). We used Hematoxylin-Eosin Staining (H&E) and Nissl staining for detecting histological changes in the spinal cord. Basso-Beattie-Bresnahan (BBB) behavioral scores were utilized for assessing functional locomotion recovery. Mouse BV2 microglia were exposed to lipopolysaccharide (LPS) to mimic SCI-induced microglial inflammation in vitro. RESULTS: Inhibition of NF-κB using JSH-23 alleviated inflammation and neuronal injury in SCI rats' spinal cords, leading to improved locomotion recovery (p < 0.05). NF-κB inhibition reduced expression levels of CD86, interleukin-6 (IL-6), IL-1ß, and inducible Nitric Oxide Synthase (iNOS), and improved expression levels of CD206, IL-4, and tissue growth factor-beta (TGF-ß) in both LPS-treated microglia and SCI rats' spinal cords (p < 0.05). Inhibition of NF-κB also effectively suppressed mitochondrial fission, evidenced by the reduced phosphorylation of dynamin-related protein 1 (DRP1) at Ser616 (p < 0.001). CONCLUSION: We show that inhibition of the NF-κB/DRP1 axis prevents mitochondrial fission and suppresses pro-inflammatory microglia polarization, promoting neurological recovery in SCI. Targeting the NF-κB/DRP1 axis therefore represents a novel approach for SCI.
Assuntos
Dinaminas , Microglia , NF-kappa B , Traumatismos da Medula Espinal , Animais , Masculino , Camundongos , Ratos , Linhagem Celular , Polaridade Celular/efeitos dos fármacos , Modelos Animais de Doenças , Dinaminas/metabolismo , Dinaminas/genética , Inflamação/metabolismo , Lipopolissacarídeos , Locomoção/efeitos dos fármacos , Microglia/metabolismo , Microglia/efeitos dos fármacos , Neuroproteção , NF-kappa B/metabolismo , Quinazolinonas , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Medula Espinal/metabolismo , Medula Espinal/efeitos dos fármacos , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/fisiopatologia , Traumatismos da Medula Espinal/patologiaRESUMO
A1 polarization of astrocytes mediated prolonged inflammation contributing to brain injury in ischemic stroke. We have previously shown that AD16 protects against neonatal hypoxic-ischemic brain damage in vivo and oxygen-glucose deprivation in vitro. More recently, AD16 has demonstrated safety, tolerability, and favorable pharmacokinetics in a randomized controlled phase I trial. In this study, we utilized a rat model of transient middle cerebral artery occlusion (tMCAO) to explore whether the anti-inflammatory compound AD16 protects against ischemic brain injury by regulating A1 polarization and its underlying mechanisms. Our results showed that AD16 treatment significantly reduced the brain infarcted volume and improved neurological function in tMCAO rats. GO analysis results show that differential genes among the Sham, tMCAO and AD16 treatment groups are involved in the regulation of cytokine and inflammatory response. KEGG enrichment pathways analysis mainly enriched in cytokine-cytokine receptor interaction, viral protein interaction with cytokine-cytokine receptor, TNF, chemokine, NF-κB and IL-17 signaling pathway. Furthermore, AD16 treatment decreased the permeability of the blood-brain barrier and suppressed neuroinflammation. AD16 treatment also significantly reduced the polarization of A1 and inhibited NF-κB and JAK2/STAT3 signaling pathways. This study demonstrates that AD16 protects against brain injury in ischemic stroke by reducing A1 polarization to suppress neuroinflammation through downregulating NF-κB and JAK2/STAT3 signaling. Our findings uncover a potential molecular mechanism for AD16 and suggest that AD16 holds promising therapeutic potential against cerebral ischemia.
Assuntos
Astrócitos , Doenças Neuroinflamatórias , Animais , Masculino , Ratos , Anti-Inflamatórios/farmacologia , Astrócitos/metabolismo , Astrócitos/efeitos dos fármacos , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Isquemia Encefálica/tratamento farmacológico , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Polaridade Celular/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Infarto da Artéria Cerebral Média/tratamento farmacológico , Infarto da Artéria Cerebral Média/patologia , Infarto da Artéria Cerebral Média/metabolismo , Doenças Neuroinflamatórias/tratamento farmacológico , Doenças Neuroinflamatórias/metabolismo , Fármacos Neuroprotetores/farmacologia , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacosRESUMO
Neuropathic pain (NP) is associated with astrocytes activation induced by nerve injury. Reactive astrocytes, strongly induced by central nervous system damage, can be classified into A1 and A2 types. Vitexin, a renowned flavonoid compound, is known for its anti-inflammatory and analgesic properties. However, its role in NP remains unexplored. This study aims to investigate the effects of vitexin on astrocyte polarization and its underlying mechanisms. A mouse model of NP was established, and primary astrocytes were stimulated with sphingosine-1-phosphate (S1P) to construct a cellular model. The results demonstrated significant activation of spinal astrocytes on days 14 and 21. Concurrently, reactive astrocytes predominantly differentiated into the A1 type. Western blot analysis revealed an increase in A1 astrocyte-associated protein (C3) and a decrease in A2 astrocyte-associated protein (S100A10). Serum S1P levels increased on days 14 and 21, alongside a significant upregulation of Sphingosine-1-phosphate receptor 1 (S1PR1) mRNA expression and elevated expression of chemokines. In vitro, stimulation with S1P inhibited the Phosphatidylinositol 3-kinase and protein kinase B (PI3K/Akt) signaling pathway and autophagy flux, promoting polarization of astrocytes towards the A1 phenotype while suppressing the polarization of A2 astrocytes. Our findings suggest that vitexin, acting on astrocytes but not microglia, attenuates S1P-induced downregulation of PI3K/Akt signaling, restores autophagy flux in astrocytes, regulates A1/A2 astrocyte ratio, and reduces chemokine and S1P secretion, thereby alleviating neuropathic pain caused by nerve injury.
Assuntos
Apigenina , Astrócitos , Autofagia , Lisofosfolipídeos , Neuralgia , Fosfatidilinositol 3-Quinases , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Receptores de Esfingosina-1-Fosfato , Esfingosina , Animais , Apigenina/farmacologia , Apigenina/uso terapêutico , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Astrócitos/patologia , Neuralgia/tratamento farmacológico , Neuralgia/metabolismo , Neuralgia/patologia , Receptores de Esfingosina-1-Fosfato/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Camundongos , Transdução de Sinais/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Lisofosfolipídeos/metabolismo , Esfingosina/análogos & derivados , Esfingosina/farmacologia , Masculino , Camundongos Endogâmicos C57BL , Polaridade Celular/efeitos dos fármacosRESUMO
Ulcerative colitis (UC) is a chronic non-specific inflammatory disease involving the mucosa and submucosa of the rectum and colon. Lindera aggregate (Sims) Kosterm is a traditional Chinese herb used for thousands of years in the treatment of gastrointestinal diseases. Previously, we have demonstrated that the extracts of Lindera aggregate have good anti-UC effects, but their pharmacodynamic active components have not been fully clarified. Therefore, we explored the therapeutic effect of Linderanine C (LDC), a characteristic component of Lindera aggregata, on UC and its mechanism in this study. Firstly, we found that LDC could significantly reduce the disease activity index of UC and improve shortened colon and pathological changes in vivo. Colon tissue transcriptomics suggested that the anti-UC effect of LDC might be related to its anti-inflammatory activity. Cellular experiments revealed that LDC could inhibit the expression of the M1 cell marker CD86 in RAW264.7 cells, reduce the production of inflammatory mediators such as IL-6 and TNF-α, and have good anti-inflammatory activity in vitro. Cellular transcriptomics reveal the potential involvement of the MAPK signaling pathway in the anti-inflammatory effect of LDC. The co-culture assay confirmed that LDC could significantly reduce inflammation-mediated intestinal epithelial cell injury. In conclusion, LDC was able to inhibit macrophage M1 polarization and reduce inflammatory mediator production by inhibiting the MAPK signaling pathway, effectively improving UC.
Assuntos
Anti-Inflamatórios , Colite Ulcerativa , Sistema de Sinalização das MAP Quinases , Macrófagos , Animais , Colite Ulcerativa/tratamento farmacológico , Colite Ulcerativa/patologia , Colite Ulcerativa/metabolismo , Camundongos , Células RAW 264.7 , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Anti-Inflamatórios/farmacologia , Colo/efeitos dos fármacos , Colo/patologia , Colo/metabolismo , Camundongos Endogâmicos C57BL , Humanos , Polaridade Celular/efeitos dos fármacos , Mediadores da Inflamação/metabolismo , Modelos Animais de DoençasRESUMO
Our understanding of how fluid forces influence cell migration in confining environments remains limited. By integrating microfluidics with live-cell imaging, we demonstrate that cells in tightly-but not moderately-confined spaces reverse direction and move upstream upon exposure to fluid forces. This fluid force-induced directional change occurs less frequently when cells display diminished mechanosensitivity, experience elevated hydraulic resistance, or sense a chemical gradient. Cell reversal requires actin polymerization to the new cell front, as shown mathematically and experimentally. Actin polymerization is necessary for the fluid force-induced activation of NHE1, which cooperates with calcium to induce upstream migration. Calcium levels increase downstream, mirroring the subcellular distribution of myosin IIA, whose activation enhances upstream migration. Reduced lamin A/C levels promote downstream migration of metastatic tumor cells by preventing cell polarity establishment and intracellular calcium rise. This mechanism could allow cancer cells to evade high-pressure environments, such as the primary tumor.