RESUMO
OBJECTIVES: Endothelial-to-mesenchymal transition (EndoMT) is a significant biological phenomenon wherein endothelial cells undergo a loss of their endothelial traits and progressively acquire mesenchymal characteristics. Consequently, this transformation leads to both a compromised ability to maintain lumen permeability and alterations in vascular structure, which hampers the preservation of blood-brain barrier integrity. This study aimed to investigate inflammation-induced EndoMT and its etiology, with the goal of impeding the infiltration of peripheral inflammation into the central nervous system. MATERIALS AND METHODS: Lipolysaccharide (LPS) was administered intraperitoneally to mice several times to establish a chronic inflammatory model. A cellular inflammatory model was established by LPS in human brain microvascular endothelial cells (HBMECs). The mRNA expressions of inflammatory cytokines interleukin-1ß (IL-1ß) and IL-6 were detected by real-time polymerase chain reaction (PCR). Immunofluorescence staining of platelet endothelial cell adhesion molecule-1 (CD31) and alpha smooth muscle actin (α-SMA) was conducted to assess the level of EndoMT. The expression levels of Occludin, zona occludens protein 1 (ZO-1), Sestrin2, microtubule-associated protein1 light chain 3 (LC3) and inducible nitric oxide synthase (iNOS) were detected by western blotting. RESULTS: LPS treatment induced the downregulation of ZO-1 and Occludin, which was accompanied by the elevated expressions of iNOS, α-SMA, Sestrin2 and LC3-II in the mouse cortex and HBMECs. Mechanistically, the knockdown of Sestrin2 in HBMECs exacerbated the EndoMT induced by LPS treatment, while the overexpression of Sestrin2 inhibited this process. Moreover, the induction of autophagy by rapamycin rescued the EndoMT induced by Sestrin2 knockdown. CONCLUSION: This study revealed that Sestrin2 inhibited endothelial inflammation and EndoMT via enhanced autophagy, which may provide a potential drug target for cerebrovascular inflammatory injury.
Assuntos
Autofagia , Células Endoteliais , Lipopolissacarídeos , Animais , Lipopolissacarídeos/farmacologia , Lipopolissacarídeos/administração & dosagem , Camundongos , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Humanos , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Camundongos Endogâmicos C57BL , Masculino , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Transição Epitelial-Mesenquimal/fisiologia , Proteínas Nucleares/metabolismo , Inflamação/metabolismo , Modelos Animais de DoençasRESUMO
The study aimed to investigate the therapeutic potential of 1,1-dimethyl-4-phenylpiperazinium iodide (DMPP), an agonist of nicotinic acetylcholine receptor (nAChR), in treating acute lung injury (ALI) induced by lipopolysaccharide (LPS). A murine ALI model was developed utilizing intraperitoneal injection of LPS. We evaluated the therapeutic efficacy of DMPP treatment in LPS-induced lung injury using various approaches, including pathohistological evaluation, appraisal of pulmonary edema, and measurement of inflammatory cytokine levels and their associated pathways within lung tissues. The gene chip data of LPS-induced acute lung injury mice were retrieved from the Gene Expression Omnibus (GEO) database for gene differential expression analysis and Gene Set Enrichment Analysis (GSEA) analysis. The impact of DMPP on glycocalyx shedding was assessed by measuring the expression levels of syndecan-1 (SDC-1) and matrix metalloproteinase-9 (MMP-9). DMPP treatment significantly improved pathomorphological changes and pathological lung injury scores in the LPS-induced ALI mouse model. The genes expressed differentially in the LPS-induced ALI group in GSE2411 were found to be involved in multiple processes, including the NF-κB signaling pathway, NOD-like receptor signaling pathway, Toll-like receptor signaling pathway, as well as the JAK-STAT signaling pathway. DMPP treatment effectively downregulated pro-inflammatory cytokines, suppressed the NF-κB signaling pathway, and effectively restrained the LPS-induced upregulation of MMP-9 and shedding of syndecan-1, thereby contributing to the preservation of endothelial glycocalyx and attenuation of endothelial barrier dysfunction. The administration of DMPP has been shown to confer protection against LPS-induced acute lung injury via a cholinergic anti-inflammatory pathway, which effectively inhibits endothelial glycocalyx degradation.
Assuntos
Lesão Pulmonar Aguda , NF-kappa B , Camundongos , Animais , NF-kappa B/metabolismo , Lipopolissacarídeos/toxicidade , Metaloproteinase 9 da Matriz/metabolismo , Sindecana-1/efeitos adversos , Iodeto de Dimetilfenilpiperazina/uso terapêutico , Iodetos/efeitos adversos , Glicocálix/metabolismo , Neuroimunomodulação , Lesão Pulmonar Aguda/tratamento farmacológico , Citocinas/efeitos adversos , Citocinas/metabolismoRESUMO
A disintegrin and metalloproteinases (ADAMs) are involved in multiple neurodegenerative diseases. However, the roles and mechanisms of ADAMs in HIV-associated neurocognitive disorder (HAND) remain unclear. Transactivator of transcription (Tat) induces inflammatory response in astrocytes, thereby leading to neuronal apoptosis in the central nervous system. In this study, we determined that ADAM17 expression was upregulated during soluble Tat stimulus in HEB astroglial cells. Inhibition of ADAM17 suppressed Tat-induced pro-inflammatory cytokines production and rescued the astrocytes-derived conditioned media (ACM)-mediated SH-SY5Y neural cells apoptosis. Moreover, ADAM17 mediated Tat-triggered inflammatory response in a NF-κB-dependent manner. Conversely, Tat induced ADAM17 expression via NF-κB signaling pathway. In addition, pharmacological inhibition of NF-κB signaling inhibited Tat-induced inflammatory response, which could be rescued by overexpression of ADAM17. Taken together, our study clarifies the potential role of the ADAM17/NF-κB feedback loop in Tat-induced inflammatory response in astrocytes and the ACM-mediated neuronal death, which could be a novel therapeutic target for relief of HAND.
Assuntos
HIV-1 , Neuroblastoma , Humanos , NF-kappa B/genética , NF-kappa B/metabolismo , HIV-1/metabolismo , Astrócitos/metabolismo , Transativadores/metabolismo , Retroalimentação , Neuroblastoma/metabolismo , Produtos do Gene tat do Vírus da Imunodeficiência Humana/metabolismo , Proteína ADAM17/genética , Proteína ADAM17/metabolismoRESUMO
Fms-like tyrosine kinase 3 (FLT3) is a glycoprotein, that is a member of the class III receptor tyrosine kinase family. Approximately one-third of acute myeloid leukemia (AML) patients have mutations of this gene, and activation of the FLT3 downstream pathway plays an important role in both normal and malignant hematopoiesis. However, the role of N-glycosylation for FLT3 activation remains unclear. In this study, we showed that the N-glycan structures on wild type (WT), internal tandem duplication (ITD), and tyrosine kinase domain (TKD) mutants of FLT3 were different. Interestingly, expression of either WT or mutant FLT3 in Ba/F3 cells, an interleukin-3 (IL-3)-dependent hematopoietic progenitor cell, greatly induced core fucosylation. To elucidate the function of core fucosylation in FLT3-mediated signaling, we used a CRISPR/Cas9 system to establish α1,6-fucosyltransferase (Fut8) knockout (KO) cells. Surprisingly, the Fut8KO resulted in cell proliferation in an IL-3-independent manner in FLT3-WT cells, which was not observed in the parental cells, and suggested that this proliferation is dependent on FLT3 expression. Fut8KO greatly increased cellular tyrosine phosphorylation levels, together with an activation of STAT5, AKT, and ERK signaling, which could be completely neutralized by restoration with Fut8 in the KO cells. Consistently, a tyrosine kinase inhibitor efficiently inhibited cell proliferation induced by Fut8KO or specific fucosylation inhibitor. Additionally, immunostaining with FLT3 showed that the proteins were mainly expressed on the cell surface in the KO cells, which is similar to FLT3-WT cells, but different from the ITD mutant. Finally, we found that Fut8KO could induce dimer-formation in FLT3 without ligand-stimulation. Taken together, the present study clearly defines the regulatory function of core fucosylation in FLT3, which could provide a valuable direction for development of drugs could be effective in the treatment of AML.
Assuntos
Fucose/metabolismo , Transdução de Sinais , Tirosina Quinase 3 Semelhante a fms/metabolismo , Animais , Glicosilação , Células HEK293 , Humanos , Interleucina-3/metabolismo , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Mutação , Domínios Proteicos , Multimerização Proteica , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT5/metabolismo , Tirosina Quinase 3 Semelhante a fms/química , Tirosina Quinase 3 Semelhante a fms/genéticaRESUMO
The α2,3-sialylation of N-glycans is considered important but complicated because the functions of the three ß-galactoside α2,3-sialyltransferases, ST3GAL3, ST3GAL4, and ST3GAL6, could be compensating for one another. To distinguish their specific functions, we established each individual knockout (KO) cell line. Loss of either the ST3GAL3 or ST3GAL6 genes decreased cell proliferation and colony formation, as opposed to the effect in the ST3GAL4 KO cells. The phosphorylation levels of ERK and AKT were significantly suppressed in the ST3GAL6 KO and ST3GAL3 KO cells, respectively. The cell aggregations were clearly observed in the KO cells, particularly the ST3GAL3 KO and ST3GAL6 KO cells, and the expression levels of E-cadherin and claudin-1 were enhanced in both those cell lines, but were suppressed in the ST3GAL4 KO cells. Those alterations were reversed with an overexpression of each corresponding gene in rescued cells. Of particular interest, the α2,3-sialylation levels of ß1 integrin were clearly suppressed in the ST3GAL4 KO cells, but these were increased in the ST3GAL3 KO and ST3GAL6 KO cells, whereas the α2,3-sialylation levels of EGFR were significantly decreased in the ST3GAL6 KO cells. The decrease in α2,3-sialylation increased the α2,6-sialylation on ß1, but not EGFR. Furthermore, a cross-restoration of each of the three genes in ST3GAL6 KO cells showed that overexpression of ST3GAL6 sufficiently rescued the total α2,3-sialylation levels, cell morphology, and α2,3-sialylation of EGFR, whereas the α2,3-sialylation levels of ß1 were greatly enhanced by an overexpression of ST3GAL4. These results clearly demonstrate that the three α2,3-sialyltransferases modify characteristic target proteins and regulate cell biological functions in different ways.
Assuntos
Sialiltransferases/metabolismo , Caderinas/metabolismo , Linhagem Celular , Proliferação de Células/fisiologia , Glicosilação , Humanos , Fosforilação/fisiologia , Transporte Proteico , Sialiltransferases/genética , beta-Galactosídeo alfa-2,3-SialiltransferaseRESUMO
The epithelial cell adhesion molecule (EpCAM) is one of the most frequently and intensely expressed of tumor-associated antigens, but the role that EpCAM plays in the proliferation, adhesion and migration properties of cancer cells remains unclear. In the present study, we screened several tumor cell lines and found that colorectal cancer CW-2 and epidermoid carcinoma A431 cells expressed relatively higher levels of EpCAM. In order to assess the biological functions of EpCAM expression in cell adhesion and migration, we established a knock out (KO) of EpCAM genes in both of these types of cancer cells via a CRISPR/Cas9 system. The elongated cell morphology was converted to a rounded morphology in the EpCAM-KO cells. These cells showed decreases in cell proliferation and migration into extracellular matrix proteins, as well as decreases in cellular signaling elements such as phosphorylated focal adhesion kinase (FAK), AKT and ERK. Moreover, the cell growth and the colony formation abilities were significantly decreased in EpCAM-KO cells. Importantly, co-immunoprecipitation analysis revealed that EpCAM associated with integrin ß1. Also, the expression levels of integrin α5 were decreased in EpCAM-KO cells, compared with that in the wild-type cells. Taken together, these data clearly demonstrate that EpCAM associates with integrin ß1 to regulate FAK/ERK signaling pathways in controlling cell adhesion, migration and proliferation via extracellular matrix adhesion, which provides novel mechanisms for EpCAM-mediated biological functions and cancer phenotypes.
Assuntos
Molécula de Adesão da Célula Epitelial/metabolismo , Integrina alfa5/metabolismo , Integrina beta1/metabolismo , Neoplasias/patologia , Adesão Celular/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Humanos , Integrina alfa5/genética , Integrina beta1/genética , Neoplasias/genética , Transdução de SinaisRESUMO
Trace metal contamination of soil is an increasing problem. Organic acid application can restore trace metal elements such as cadmium (Cd) in contaminated soil. Changbai larch (Larix olgensis A. Henry) is an economically important forestry species in northeast China; however, growth is inhibited by severe Cd contamination. We investigated the effects of different concentrations of exogenous succinic acid (SA) on Cd tolerance and physiological and morphological toxicity in L. olgensis seedlings. Seedlings were planted in pots containing Cd-contaminated or uncontaminated Haplic Cambisol. Seedlings in Cd-contaminated soil were treated daily with SA solution at 0, 0.04, 0.2, 1.0, and 2.0 mmol kg-1 of soil for 10, 20 or 30 days. Cd treatment induced seedling damage and significantly increased the relative conductivity and malondialdehyde content of the leaves, inhibiting soluble protein and proline contents, superoxide dismutase and peroxidase activity, chlorophyl fluorescence and pigment content. Decreases in the length, surface area, volume of roots and leaves, and specific root length were also observed. Effects increased in control plants with time. SA treatment also reduced the Cd content of the fine roots and leaves and Mg, K, and Ca contents. Moreover, plant growth was significantly promoted and damage was reversed, especially at 5.0 and 10.0 mmol L-1 SA for 30 days. SA therefore alleviated Cd-induced injury, improving tolerance to Cd stress. SA application combined with afforestation could therefore help restore Cd-contaminated soil in northeast China. Further studies aimed at determining the detoxification mechanism of L. olgensis seedlings are now required.
Assuntos
Cádmio , Larix , Biodegradação Ambiental , China , Folhas de Planta , Raízes de Plantas , Plântula , Ácido SuccínicoRESUMO
Mixed Pb and Cd soil contamination is an issue in Northeast China. We examined the effects of exogenous organic acids on the resilience of Changbai larch (Larix olgensis) seedlings, a pioneering forestry species in afforestation and vegetation restoration in Northeast China, under such stress. Mixed Pb and Cd stress led to significantly higher Pb and Cd content in the leaves and fine roots, malondialdehyde content in the leaves, superoxide dismutase activity, and soluble protein content in the leaves. Lower biomass of the roots, stems, and leaves was observed, with the roots showing the sharpest reduction in biomass. However, the application of organic acids mitigated or reversed these effects. This was most pronounced following treatment with 0.2 mmol·L-1 or 1.0 mmol·L-1 organic acids for 20 days. Citric acid had the greatest positive effect compared with succinic acid and oxalic acid. We suggest that exogenous organic acids have the potential to alleviate Pb and Cd-induced oxidation injury symptoms in Changbai larch seedlings, and may enhance resilience to mixed Pb and Cd stress.
RESUMO
DIX domain containing 1 (Dixdc1), a positive regulator of Wnt signaling pathway, is recently reported to play a role in the neurogenesis. However, the distribution and function of Dixdc1 in the central nervous system (CNS) after brain injury are still unclear. We used an acute traumatic brain injury (TBI) model in adult rats to investigate whether Dixdc1 is involved in CNS injury and repair. Western blot analysis and immunohistochemistry showed a time-dependent up-regulation of Dixdc1 expression in ipsilateral cortex after TBI. Double immunofluorescent staining indicated a colocalization of Dixdc1 with astrocytes and neurons. Moreover, we detected a colocalization of Ki-67, a cell proliferation marker with GFAP and Dixdc1 after TBI. In primary cultured astrocytes stimulated with lipopolysaccharide, we found enhanced expression of Dixdc1 in parallel with up-regulation of Ki-67 and cyclin A, another cell proliferation marker. In addition, knockdown of Dixdc1 expression in primary astrocytes with Dixdc1-specific siRNA transfection induced G0/G1 arrest of cell cycle and significantly decreased cell proliferation. In conclusion, all these data suggest that up-regulation of Dixdc1 protein expression is potentially involved in astrocyte proliferation after traumatic brain injury in the rat.
Assuntos
Astrócitos/metabolismo , Astrócitos/patologia , Lesões Encefálicas Traumáticas/metabolismo , Lesões Encefálicas Traumáticas/patologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Animais , Proliferação de Células , Células Cultivadas , Córtex Cerebral/metabolismo , Córtex Cerebral/patologia , Imuno-Histoquímica , Antígeno Ki-67/metabolismo , Masculino , Neurônios/metabolismo , Neurônios/patologia , Ratos Sprague-Dawley , Regulação para CimaRESUMO
Ischemic stroke is a dominant health problem with extremely high rates of mortality and disability. The main mechanism of neuronal injury after stroke is excitotoxicity, during which the activation of neuronal nitric oxide synthase (nNOS) exerts a vital role. However, directly blocking N-methyl-D-aspartate receptors or nNOS can lead to severe undesirable effects since they have crucial physiological functions in the central nervous system. Here, we report that nNOS undergoes O-linked-ß-N-acetylglucosamine (O-GlcNAc) modification via interacting with O-GlcNAc transferase, and the O-GlcNAcylation of nNOS remarkably increases during glutamate-induced excitotoxicity. In addition, eliminating the O-GlcNAcylation of nNOS protects neurons from apoptosis during glutamate stimulation by decreasing the formation of nNOS-postsynaptic density protein 95 complexes. Taken together, our data suggest a novel function of the O-GlcNAcylation of nNOS in neuronal apoptosis during glutamate excitotoxicity, suggesting a novel therapy strategy for ischemic stroke.
Assuntos
Acetilglucosamina/metabolismo , Apoptose/fisiologia , Ácido Glutâmico/toxicidade , N-Acetilglucosaminiltransferases/metabolismo , Neurônios/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Animais , Apoptose/efeitos dos fármacos , Glicosilação/efeitos dos fármacos , Masculino , Neurônios/efeitos dos fármacos , Células PC12 , Ratos , Ratos Sprague-DawleyRESUMO
The insulin-like growth factor (IGF) system is linked to CNS pathological states. The functions of IGFs are modulated by a family of binding proteins termed insulin-like growth factor binding proteins (IGFBPs). Here, we demonstrate that IGFBP-6 may be associated with neuronal apoptosis in the processes of intracerebral hemorrhage (ICH). We obtained a significant upregulation of IGFBP-6 in neurons adjacent to the hematoma following ICH with the results of Western blot, immunohistochemistry, and immunofluorescence. Increasing IGFBP-6 level was found to be accompanied by the upregulation of Bax, Bcl-2, and active caspase-3. Besides, IGFBP-6 co-localized well with active caspase-3 in neurons, indicating its potential role in neuronal apoptosis. Knocking down IGFBP-6 by RNA-interference in PC12 cells reduced active caspase-3 expression. Thus, IGFBP-6 may play a role in promoting the brain secondary damage following ICH.
Assuntos
Apoptose/fisiologia , Hemorragia Cerebral/metabolismo , Proteína 6 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Neurônios/metabolismo , Animais , Hemorragia Cerebral/patologia , Masculino , Neurônios/patologia , Células PC12 , Ratos , Ratos Sprague-DawleyRESUMO
Ring finger protein 1 (RING1) is a RING domain characterized protein belonging to the RING finger family. It is an E3 ubiquitin-protein ligase that mediated monoubiquitination of histone H2A and the core component of PRC1 complex, which is the repressive multiprotein complex of Polycomb group (PcG). Previous studies showed the important tumorigenic role of RING1 via promoting cell proliferation and the crucial function in maintaining transcriptional program stability during development. However, its mechanism for spinal cord injury (SCI) is still unknown. In our research, we established an acute SCI model in adult rats and studied the expression and function profiles of RING1. RING1 protein level detected by western blot peaked at day 3 after trauma and then decreased gradually. Immunohistochemistry showed the increase of RING1 expression displayed in the white matter more obviously than in the gray matter. Furthermore, increased co-expression of RING1 and GFAP confirmed activated astrocytes in injured spinal cord via double immunofluorescence staining. Meanwhile, we also found the co-localization of PCNA, a famous marker of proliferative cells, with RING1 and GFAP, which indicated RING1 might play a role in astrocyte proliferation after SCI. In vitro studies, RING1 protein level in C6 cells increased after LPS challenge and RING1 was required for astrocyte proliferation and activation induced by LPS. In summary, we took a new insight into the function of RING1 in the cellular and molecular mechanism underlying the pathophysiology of SCI.
Assuntos
Neuroglia/metabolismo , Complexo Repressor Polycomb 1/biossíntese , Traumatismos da Medula Espinal/metabolismo , Traumatismos da Medula Espinal/patologia , Ubiquitina-Proteína Ligases/biossíntese , Animais , Expressão Gênica , Masculino , Neuroglia/patologia , Complexo Repressor Polycomb 1/genética , Ratos , Ratos Sprague-Dawley , Medula Espinal/citologia , Medula Espinal/metabolismo , Medula Espinal/patologia , Traumatismos da Medula Espinal/genética , Fatores de Tempo , Ubiquitina-Proteína Ligases/genéticaRESUMO
Microglial activation, including classical (M1) and alternative (M2) activation, plays important roles in the development of several central nervous system disorders and promotes tissue reconstruction. Toll-like receptor (TLR)4 is important for microglial polarization. TIR domain-containing adaptor protein (TIRAP) is an intracellular adaptor protein, which is responsible for the early phase of TLR4 activation. The role of TIRAP in BV2 cell M1 polarization is still unknown. In this study, we showed that TIRAP expression is greatly elevated in lipopolysaccharide (LPS)/interferon (IFN)-γ-treated microglia. TIRAP overexpression promoted BV2 microglial M1 polarization by increasing M1-related marker production (inducible nitric oxide synthase, CD86, interleukin-6, interleukin-1ß and tumour necrosis factor-α). In contrast, TIRAP knockdown prevented M1-related marker production. Mechanistically, TIRAP could interact with TNF Receptor-Associated Factor 6 (TRAF6) to increase M1-related marker production in TIRAP overexpressed and LPS/IFN-γ-treated BV2 cells. In addition, silencing of TIRAP effectively inhibited the activation of the Transforming Growth Factor-Beta-Activated Kinase 1/I-Kappa-B Kinase /Nuclear Factor of Kappa Light Polypeptide Gene Enhancer in B-Cells (TAK1/IKK/NF-κB) signalling pathway and the phosphorylation of Akt and mitogen-activated protein kinases, which were activated by LPS/IFN-γ stimulation. Thus, our results suggest that TIRAP positively regulated BV2 microglial M1 polarization through TLR4-mediated TAK1/IKK/NF-κB, mitogen-activated protein kinases and Akt signalling pathways.
Assuntos
Polaridade Celular , Glicoproteínas de Membrana/metabolismo , Microglia/metabolismo , Receptores de Interleucina-1/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Encefalite/induzido quimicamente , Encefalite/metabolismo , Quinase I-kappa B/metabolismo , Interferon gama/administração & dosagem , Lipopolissacarídeos , MAP Quinase Quinase Quinases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/metabolismo , NF-kappa B/metabolismo , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptor 4 Toll-Like/metabolismoRESUMO
Naked2 (NKD2), one member of Naked family, has been shown to negatively regulate Wnt/ß-catenin signaling pathway. It has been recognized that NKD2 plays a vital role in cell homeostasis and prevention of tumorigenesis. However, NKD2 expression and its functional role in the brain in neuroinflammatory processes remain unclear. In our study, we investigated NKD2 distribution and role in lipopolysaccharide (LPS)-induced neuroinflammation rat model. The data indicated that NKD2 was up-regulated in LPS-injected brain, and the cellular localization of NKD2 was predominantly in cerebral cortical neurons. Furthermore, we treated primary neurons with conditioned media (CM) collected from LPS-stimulated mixed glial cultures (MGC). We detected that the up-regulation of NKD2 might be associated with the subsequent apoptosis in neurons. We also found knockdown NKD2 partially depressed the increase of cleaved caspase-3 and increased the reduction of ß-catenin stimulated by MGC-CM. Taken together, these results suggested that NKD2 might be involved in neuronal apoptosis via the Wnt/ß-catenin pathway during neuroinflammation in CNS. Our findings might provide a new therapeutic target for the prevention of neuroinflammation-involved neurological disorders.
Assuntos
Proteínas de Transporte/metabolismo , Córtex Cerebral/patologia , Inflamação/patologia , Lipopolissacarídeos/farmacologia , Neurônios/patologia , Regulação para Cima/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Biomarcadores/metabolismo , Meios de Cultivo Condicionados/farmacologia , Imunofluorescência , Inativação Gênica/efeitos dos fármacos , Masculino , Ratos Sprague-Dawley , Fatores de Tempo , Via de Sinalização Wnt/efeitos dos fármacosRESUMO
Promyelocytic leukemia zinc finger (PLZF) protein has been identified as a tumor suppressor in a variety of cancers, including leukemia, malignant mesothelioma, malignant melanoma, pancreatic cancer and prostate cancer. Studies have demonstrated that altered expression of PLZF affected its biological functions associated with tumorigenesis, such as proliferation, cell cycle, and apoptosis. However, information regarding its regulation and possible function in the central nervous system diseases is still limited. In this study, we performed a neuroinflammatory model by lipopolysaccharide (LPS) lateral ventricle injection in adult rats and detected increased expression of PLZF in the brain cortex. Immunofluorescence assay indicated that PLZF was significantly increased in neurons 3 day after LPS injection, but not in astrocytes and microglia. Moreover, there was a concomitant upregulation of active caspase-3, cyclin D1, and CDK4 in vivo and vitro studies. In addition, the expression of these proteins in cortical primary neurons was inhibited after knocking down PLZF by siRNA. Collectively, all these results suggested that the upregulation of PLZF might be involved in neuronal apoptotic-like injury in neuroinflammation after LPS injection.
Assuntos
Apoptose/efeitos dos fármacos , Proteínas de Ligação a DNA/metabolismo , Inflamação/induzido quimicamente , Lipopolissacarídeos/farmacologia , Neurônios/metabolismo , Animais , Ciclo Celular/efeitos dos fármacos , Masculino , Proteína com Dedos de Zinco da Leucemia Promielocítica , RNA Interferente Pequeno/metabolismo , Ratos Sprague-Dawley , Ativação Transcricional/fisiologia , Regulação para CimaRESUMO
Interferon regulatory factor 3 (IRF3) is a member of IRF family which plays a significant role in the innate immune response, apoptosis, and oncogenesis. Mounting evidence has demonstrated that IRF3 was involved in central nervous system disease such as cerebral ischemic injury through promoting neuronal apoptosis. However, it remains unclear about the underlying mechanisms of IRF3 upon neuronal apoptosis following intracerebral hemorrhage (ICH). In the present study, we established an adult rat ICH model by injecting autologous whole blood into the right basal ganglia and evaluated their neurological deficits by behavioral tests. IRF3 protein level was up-regulated adjacent to the hematoma following ICH when compared with the sham brain cortex by western blot and immunohistochemistry. Immunofluorescent staining indicated IRF3 was mainly localized in neurons, a few in astrocytes. In addition, we also detected that IRF3 co-localized with active caspase-3 which is a neuronal apoptosis marker. Furthermore, in vitro study, knocking down IRF3 by using IRF3 interference in primary cortical neurons reduced the expression of active caspase-3 and Bax while increased Bcl-2. In conclusion, we speculated that IRF3 might exert pro-apoptotic function in neurons after ICH.
Assuntos
Apoptose/fisiologia , Astrócitos/metabolismo , Hemorragia Cerebral/metabolismo , Fator Regulador 3 de Interferon/metabolismo , Neurônios/metabolismo , Animais , Caspase 3/metabolismo , Técnicas de Silenciamento de Genes/métodos , Masculino , Ratos , Ratos Sprague-Dawley , Ativação Transcricional/fisiologia , Regulação para CimaRESUMO
Hepatocellular carcinoma (HCC) is a major type of primary liver cancer and the sixth most prevalent human malignancies worldwide. However, the molecular mechanisms underlying hepatocarcinogenesis remain unclear. For HCC patients, there is not only a lack of effective therapeutic targets but also a lack of predictive or prognostic biomarkers. In this article, we reported that TRIM32 was obviously upregulated in HCC tumor tissues and HCC cell lines. Its expression patterns were positively correlated with histological grade, tumor sizes, and HBsAg of HCC patients. TRIM32 expression was a significant predictor for the overall survival time of HCC patients. Moreover, the overexpression of TRIM32 in cells accelerated the G1-S phase transition, promoted cell proliferation rates, and induced the resistance of HCC patients to oxaliplatin. All these findings suggest that TRIM32 might play important roles in the hepatocarcinogenesis. TRIM32 could be a novel direction to explore the mechanism underlying HCC pathogenesis.
Assuntos
Carcinoma Hepatocelular , Proliferação de Células , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas , Fatores de Transcrição/biossíntese , Proteínas com Motivo Tripartido/biossíntese , Ubiquitina-Proteína Ligases/biossíntese , Adulto , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/mortalidade , Intervalo Livre de Doença , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/mortalidade , Masculino , Pessoa de Meia-Idade , Compostos Organoplatínicos/administração & dosagem , Oxaliplatina , Taxa de SobrevidaRESUMO
Somatostatins are peptide hormones that regulate diverse cellular processes, such as neurotransmission, cell proliferation, apoptosis, and endocrine signaling as well as inhibiting the release of many hormones and other secretory proteins. SSTR1 is a member of the superfamily of somatostatin receptors possessing seven-transmembrane segments. Aberrant expression of SSTR1 has been implicated in several human diseases, including pseudotumor cerebri, and oncogenic osteomalacia. In this study, we investigated a potential role of SSTR1 in the regulation of neuronal apoptosis in the course of intracerebral hemorrhage (ICH). A rat ICH model in the caudate putamen was established and subjected to behavioral tests. Western blot and immunohistochemistry indicated a remarkable up-regulation of SSTR1 expression surrounding the hematoma after ICH. Double-labeled immunofluorescence showed that SSTR1 was mostly co-localized with neurons, and was rarely distributed in activated astrocytes and microglia. Additionally, SSTR1 co-localized with active-caspase-3 and bcl-2 around the hematoma. The expression of active-caspase-3 was parallel with that of SSTR1 in a time-dependent manner. In addition, SSTR1 knockdown specifically resulted in reduced neuronal apoptosis in PC12 cells. All our findings suggested that up-regulated SSTR1 contributed to neuronal apoptosis after ICH, which was accompanied with reduced expression of bcl-2.
Assuntos
Apoptose , Hemorragia Cerebral/metabolismo , Hemorragia Cerebral/patologia , Neurônios/patologia , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Receptores de Somatostatina/metabolismo , Regulação para Cima , Envelhecimento/patologia , Animais , Apoptose/efeitos dos fármacos , Biomarcadores/metabolismo , Western Blotting , Caspase 3/metabolismo , Hemorragia Cerebral/enzimologia , Modelos Animais de Doenças , Ativação Enzimática/efeitos dos fármacos , Imunofluorescência , Hematoma/metabolismo , Hematoma/patologia , Hemina/farmacologia , Humanos , Masculino , Neurônios/efeitos dos fármacos , Neurônios/enzimologia , Células PC12 , Ratos , Ratos Sprague-Dawley , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: The persistent presence of inflammation is a recognized pathogenic mechanisms of diabetic foot ulcers (DFUs). We aimed to investigate the expression of PLIN1 in tissues from DFU patients and assess its potential association with inflammation-induced damage. METHODS: We performed transcriptome sequencing and correlation analysis of the foot skin from patients with or without DFUs. Additionally, we examined the correlation between PLIN1 and related inflammatory indicators by analyzing PLIN1 expression in tissue and serum samples and through high-glucose stimulation of keratinocytes (HaCaT cells). RESULTS: PLIN1 is upregulated in the tissue and serum from DFU patients. Additionally, PLIN1 shows a positive correlation with leukocytes, neutrophils, monocytes, C-reactive protein, and procalcitonin in the serum, as well as IL-1ß and TNF-α in the tissues. Experiments with Cells demonstrated that reduced expression of PLIN1 leads to significantly decreased expression of iNOS, IL-1ß, IL-6, IL-18, and TNF-α. PLIN1 may mediate wound inflammatory damage through the NF-κB signaling pathway. CONCLUSION: Our findings suggest that PLIN1 mediates the inflammatory damage in DFU, offering new prospects for the treatment of DFU.
Assuntos
Diabetes Mellitus , Pé Diabético , Humanos , Pé Diabético/genética , Pé Diabético/patologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Pele/patologia , Inflamação/metabolismo , Queratinócitos/metabolismo , Diabetes Mellitus/metabolismo , Perilipina-1/metabolismoRESUMO
Introduction: Neuroinflammation serves as a critical local defense mechanism against secondary brain injury following intracerebral hemorrhage (ICH), and astrocytes play a prominent role in this process. In this study, we investigated astrocytic changes during the inflammatory state after ICH to identify new targets for improving the inflammatory response. Methods: We stimulated mouse astrocytes with lipopolysaccharide (LPS) in vitro and analyzed their transcriptomes via ribonucleic acid sequencing. We created an ICH model in living organisms by injecting autologous blood. Results: RNA sequencing revealed that 2,717 genes were differentially expressed in the LPS group compared to those in the saline group, with notable enrichment of the autophagic pathway. By intersecting the 2,717 differentially expressed genes (DEGs) with autophagy-related genes, we identified 36 autophagy-related DEGs and seven hub genes. Previous studies and quantitative reverse transcription-polymerase chain reaction results confirmed the increased expression of phosphatidylinositol 3-kinase catalytic subunit type 3 (Pik3c3), AKT serine/threonine kinase 1 (Akt1), and unc-51 like autophagy activating kinase 2 (Ulk2) in astrocytes after ICH. Transcription factors and target miRNAs were identified for the final three DEGs, and 3-methyladenine and leupeptin were identified as potential therapeutic agents for ICH. Conclusion: Our findings suggest that astrocyte autophagy plays a critical role in ICH complexity, and that Pik3c3, Akt1, and Ulk2 may be potential therapeutic targets.