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1.
São Paulo; s.n; 20220601. 73 p.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1370752

ABSTRACT

O carcinoma secretório em glândula salivar é uma neoplasia recentemente descrita que tem os mesmos aspectos morfológicos, imuno-histoquímicos e genéticos do carcinoma secretório de origem mamária. O carcinoma secretório tem características celulares reminiscentes de uma célula secretora lactacional, isto é, um citoplasma vacuolado repleto de gotas lipídicas e um material secretado, por vezes de forma apócrina, que pode lembrar o leite. Mais recentemente, algum nível de diferenciação lactacional foi sugerida no carcinoma secretório de origem salivar. O objetivo do estudo foi verificar se existe uma diferenciação do tipo lactacional em carcinomas secretórios de origem salivar, comparando a outros tipos de carcinomas salivares mais comuns. Foram realizadas reações imuno-histoquímicas para as seguintes proteínas: receptores hormonais (receptor de prolactina e receptor do hormônio do crescimento), proteínas associadas ao produto de secreção da glândula mamária lactacional (mucina-1 (MUC-1), MUC4, globulina de gordura 1 do leite humano, lactoferrina) e proteínas associadas à via Akt-mTOR (PTEN, p-Akt, p-mTOR, p4EBP1, eIF4E, pS6). A maioria dos casos de carcinoma secretório foi negativa para receptor de prolactina e de hormônio do crescimento. Lactoferrina foi positiva em todos os grupos tumorais, porém somente em carcinoma secretório observou-se um padrão de marcação intensa, difuso tanto em célula como em secreção. Todos os casos de carcinoma secretório foram positivos para globulina de gordura do tipo 1, porém o mesmo padrão de marcação foi observado em outros tumores. A maioria dos casos de carcinoma secretório foram positivos para MUC1 e MUC4. Nenhum caso de carcinoma secretório foi positivo para Akt, mas PTEN foi difusamente expresso em 57,1% dos casos. mTOR foi expresso em mais da metade dos casos de carcinoma secretório e dos outros tumores salivares. Entre as proteínas à jusante de mTOR, somente eIF4E demonstrou alta expressão no grupo de estudo. A expressão de marcadores lactacionais não é exclusiva do carcinoma secretório, porém a expressão de lactoferrina é distinta neste grupo de tumores quando comparado aos demais tumores salivares estudados.


Subject(s)
Salivary Proteins and Peptides , Lactation , Salivary Gland Neoplasms , Signal Transduction
2.
São Paulo; s.n; 20220601. 73 p.
Thesis in Portuguese | LILACS, BBO | ID: biblio-1370757

ABSTRACT

O carcinoma secretório em glândula salivar é uma neoplasia recentemente descrita que tem os mesmos aspectos morfológicos, imuno-histoquímicos e genéticos do carcinoma secretório de origem mamária. O carcinoma secretório tem características celulares reminiscentes de uma célula secretora lactacional, isto é, um citoplasma vacuolado repleto de gotas lipídicas e um material secretado, por vezes de forma apócrina, que pode lembrar o leite. Mais recentemente, algum nível de diferenciação lactacional foi sugerida no carcinoma secretório de origem salivar. O objetivo do estudo foi verificar se existe uma diferenciação do tipo lactacional em carcinomas secretórios de origem salivar, comparando a outros tipos de carcinomas salivares mais comuns. Foram realizadas reações imuno-histoquímicas para as seguintes proteínas: receptores hormonais (receptor de prolactina e receptor do hormônio do crescimento), proteínas associadas ao produto de secreção da glândula mamária lactacional (mucina-1 (MUC-1), MUC4, globulina de gordura 1 do leite humano, lactoferrina) e proteínas associadas à via Akt-mTOR (PTEN, p-Akt, p-mTOR, p4EBP1, eIF4E, pS6). A maioria dos casos de carcinoma secretório foi negativa para receptor de prolactina e de hormônio do crescimento. Lactoferrina foi positiva em todos os grupos tumorais, porém somente em carcinoma secretório observou-se um padrão de marcação intensa, difuso tanto em célula como em secreção. Todos os casos de carcinoma secretório foram positivos para globulina de gordura do tipo 1, porém o mesmo padrão de marcação foi observado em outros tumores. A maioria dos casos de carcinoma secretório foram positivos para MUC1 e MUC4. Nenhum caso de carcinoma secretório foi positivo para Akt, mas PTEN foi difusamente expresso em 57,1% dos casos. mTOR foi expresso em mais da metade dos casos de carcinoma secretório e dos outros tumores salivares. Entre as proteínas à jusante de mTOR, somente eIF4E demonstrou alta expressão no grupo de estudo. A expressão de marcadores lactacionais não é exclusiva do carcinoma secretório, porém a expressão de lactoferrina é distinta neste grupo de tumores quando comparado aos demais tumores salivares estudados.


Subject(s)
Salivary Proteins and Peptides , Lactation , Signal Transduction
3.
Braz. j. biol ; 82: e232525, 2022. tab, graf
Article in English | LILACS | ID: biblio-1249239

ABSTRACT

Abstract The coupling of a ligand with a molecular receptor induces a signal that travels through the receptor, reaching the internal domain and triggering a response cascade. In previous work on T-cell receptors and their coupling with foreign antigens, we observed the presence of planar molecular patterns able to generate electromagnetic fields within the proteins. These planes showed a coherent (synchronized) behavior, replicating immediately in the intracellular domain that which occurred in the extracellular domain as the ligand was coupled. In the present study, we examined this molecular transduction - the capacity of the coupling signal to penetrate deep inside the receptor molecule and induce a response. We verified the presence of synchronized behavior in diverse receptor-ligand systems. To appreciate this diversity, we present four biochemically different systems - TCR-peptide, calcium pump-ADP, haemoglobin-oxygen, and gp120-CD4 viral coupling. The confirmation of synchronized molecular transduction in each of these systems suggests that the proposed mechanism would occur in all biochemical receptor-ligand systems.


Resumo A ligação de um ligante com um receptor molecular induz um sinal que viaja através do receptor, chegando ao domínio interno e disparando uma cascata de resposta. Em trabalhos anteriores em receptores de células T e sua ligação com antígenos estranhos, observamos a presença de padrões moleculares planares capazes de gerar campos eletromagnéticos dentro das proteínas. Esses planos mostraram um comportamento coerente (sincronizado), replicando, instantaneamente, no domínio intracelular o que ocorreu no domínio extracelular, enquanto o ligante era acoplado. No presente estudo, examinamos essa transdução - a capacidade de um sinal de acoplamento de penetrar profundamente a molécula receptora e induzir uma resposta. Verificamos a presença de um comportamento coerente em sistemas diversos de receptor-ligante. Para apreciar essa diversidade, apresentamos quatro sistemas bioquímicos diferentes: TCR-peptídeo, ADP-bomba de cálcio, hemoglobina-oxigênio e gp120-CD4 acoplamento viral. A confirmação de transdução molecular sincronizada em cada um desses sistemas sugere que o mecanismo proposto ocorreria em todos os sistemas bioquímicos receptor-ligante.


Subject(s)
Signal Transduction , Electromagnetic Fields , Receptors, Antigen, T-Cell/genetics , Ligands
4.
São Paulo; s.n; 2022. 138 p. ilus, tab.
Thesis in Portuguese | LILACS, Inca | ID: biblio-1378924

ABSTRACT

O carcinoma renal de células claras (CRCC) é o tipo de neoplasia renal com maior incidência, cerca de 80%. A maioria dos casos são curados após cirurgia, porém, cerca de um terço dos pacientes apresentam recidiva da doença com metástase à distância. O tratamento para este tumor evoluiu muito nas últimas duas décadas, entretanto, pacientes metastáticos ainda apresentam baixas taxas de resposta aos tratamentos devido a resistência adquirida pelo tumor para escapar da terapia alvo. Identificar os mecanismos moleculares associados à carcinogênese do CRCC é essencial para entender as características tumorais que estão associadas a progressão da doença e resistência aos tratamentos. Entre as alterações mais frequentes no CRCC está a perda do gene VHL, um supressor tumoral e principal regulador da resposta à hipóxia. VHL tem dois principais alvos, o fator induzido por hipóxia 1α (HIF-1α) e o fator induzido por hipóxia α (HIF-2α). Em normóxia, VHL é responsável pela degradação das subunidades de HIF. Em hipóxia, VHL deixa de reconhecer e marcar HIF-1α e HIF-2α para degradação e, uma vez estabilizadas, ativam vias de sinalização associadas a sobrevivência celular. As informações sobre alterações encontradas em tumores normalmente são estudadas a partir do sequenciamento da população total de mRNAs, oferecendo uma visão do transcriptoma. Nossa abordagem metodológica coleta e analisa apenas a população de mRNAs ativamente traduzidos, oferecendo uma visão mais próxima da expressão proteica final. A via de mTOR regula o início da tradução de mRNAs e está frequentemente mutada em CRCC. A hipóxia afeta a expressão de genes tanto via transcrição quanto via tradução. Alterações no controle traducional em CRCC afetam a expressão gênica contribuindo para a formação do tumor e progressão da doença. Assim, nosso objetivo principal foi identificar o perfil de genes diferencialmente traduzidos dependendo do status de VHL e da via de mTOR. Para isso utilizamos um modelo celular de CRCC deficiente em VHL e sua contraparte onde VHL foi restituído. Realizamos o perfil polissomal em modelos celulares de CRCC para separar e coletar a população de mRNAs ativamente traduzidos que foram posteriormente sequenciados. Nossos dados mostraram perfis distintos de tradução entre as células VHL- deficientes e VHL-proficientes. Além disso, após a inibição de mTOR, ambas as células também apresentaram respostas diferentes ao tratamento. Além disso, observamos alterações na resposta imune e aumento do ciclo celular na ausência de VHL, que podem contribuir para a progressão tumoral. Em modelo com tecido tumoral congelado, nossos resultados parciais indicam que alterações na tradução global podem interferir principalmente no estadiamento clínico de pacientes com CRCC. Por fim, também analisamos a expressão de HIF-2α, um dos alvos de VHL, em tecidos de pacientes com CRCC. Nossos resultados mostram que HIF-2α pode ser utilizado na estratificação de pacientes com maior risco de recidiva, dependendo do estadiamento clínico.


Clear cell renal cell carcinoma (ccRCC) is the most common type of renal neoplasia with 80% of incidence. Most cases are cured after surgery, however, one third of all patients will have disease recurrence with distant metastasis. ccRCC treatment had evolved in the past two decades, however, metastatic patients still have low response rates due to tumor resistance. The identification of molecular mechanisms associated with ccRCC carcinogenesis is essential to understand the characteristics associated with disease progression and treatment resistance. The most frequent alteration in ccRCC is the loss of VHL gene, a tumor suppressor and the main regulator in response to hypoxia. VHL has two main target, hypoxia-induced factor 1 α (HIF-1 α) and hypoxia-induced factor α (HIF-2 α). In normoxic conditions, VHL can lead HIF subunits to degradation. In hypoxia, HIF-1α and HIF-2α stabilize and activate cell survival associated signaling pathways. Studies about tumor alterations usually provides a view of the transcriptome. Our approach is based on the actively translated mRNAs collection and analysis, which provides a closer view from protein expression. mTOR pathway regulates translation initiation and is frequently mutated in ccRCC. Hypoxia affects gene expression in both transcriptional and translational regulation. Alteration in translational control in ccRCC affect gene expression which contributes to tumor progression. Our main objective was to identify the differentially translated gene profile depending on VHL status and mTOR pathway activation. To assess this, we used a VHL-deficient and a VHL-proficient ccRCC cell line. We used the polysome profiling technique to separate and collect the population of mRNAs actively translated that were subsequently sequenced. Our data showed distinct translation profiles between VHL-deficient and VHL-proficient cells. In addition, after mTOR inhibition, both cells showed different responses to treatment. We observed changes in immune response and increased cell cycle pathways in VHL deficient cells, which may contribute to tumor progression. In tumor tissue, our polysome profiling analysis indicate that changes in global translation may interfere in clinical staging of ccRCC patients. Finally, we analyzed the expression of HIF-2α, a VHL target, in ccRCC patient's tissues. Our results showed that HIF-2α can distinct patients at higher recurrence risk depending on clinical staging.


Subject(s)
Humans , RNA, Messenger/genetics , Carcinoma, Renal Cell/genetics , Gene Expression Profiling , Von Hippel-Lindau Tumor Suppressor Protein , Kidney Neoplasms/genetics , Signal Transduction , Gene Expression Regulation, Neoplastic
5.
Article in Chinese | WPRIM | ID: wpr-936368

ABSTRACT

OBJECTIVE@#To explore the therapeutic mechanism of tanshinone IIA in the treatment of pulmonary arterial hypertension (PAH) in rats.@*METHODS@#A total of 100 male SD rats were randomized into 5 groups (n=20), and except for those in the control group with saline injection, all the rats were injected with monocrotaline (MCT) on the back of the neck to establish models of pulmonary hypertension. Two weeks after the injection, the rat models received intraperitoneal injections of tanshinone IIA (10 mg/kg), phosphatidylinositol 3 kinase (PI3K) inhibitor (1 mg/kg), both tanshinone IIA and PI3K inhibitor, or saline (model group) on a daily basis. After 2 weeks of treatment, HE staining and α-SMA immunofluorescence staining were used to evaluate the morphology of the pulmonary vessels of the rats. The phosphorylation levels of PI3K, protein kinase B (PKB/Akt) and endothelial nitric oxide synthase (eNOS) in the lung tissue were determined with Western blotting; the levels of eNOS and NO were measured using enzyme-linked immunosorbent assay (ELISA).@*RESULTS@#The results of HE staining and α-SMA immunofluorescence staining showed that tanshinone IIA effectively inhibited MCT-induced pulmonary artery intimamedia thickening and muscularization of the pulmonary arterioles (P < 0.01). The results of Western blotting showed that treatment with tanshinone IIA significantly increased the phosphorylation levels of PI3K, Akt and eNOS proteins in the lung tissue of PAH rats; ELISA results showed that the levels of eNOS and NO were significantly decreased in the rat models after tanshinone IIA treatment (P < 0.01).@*CONCLUSION@#Treatment with tanshinone IIA can improve MCT-induced pulmonary hypertension in rats through the PI3K/Akt-eNOS signaling pathway.


Subject(s)
Abietanes , Animals , Hypertension, Pulmonary/drug therapy , Male , Monocrotaline/toxicity , Nitric Oxide Synthase Type III/therapeutic use , Phosphatidylinositol 3-Kinase/pharmacology , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Pulmonary Artery , Rats , Rats, Sprague-Dawley , Signal Transduction
6.
Article in Chinese | WPRIM | ID: wpr-936361

ABSTRACT

OBJECTIVE@#To investigate the role of proline 4-hydroxylase Ⅱ (P4HA2) in the occurrence and progression of liver cancer.@*METHODS@#GEPIA and Human Protein Atlas database were used to predict the expression of P4HA2 in hepatocellular carcinoma (HCC), and K-M plotter online database was used to analyze the relationship between P4HA2 expression and the prognosis of HCC. We also examined the expressions of P4HA2 in HCC cells and normal hepatocytes using qRT-PCR and Western blotting. With lentivirus-mediated RNA interference, P4HA2 expression was knocked down in hepatoma SNU-449 and Hep-3B cells, and the changes in cell proliferation, migration and invasion were assessed using cell counting kit-8 (CCK-8) assay, colony formation test, scratch test and Transwell assay. The changes in the expressions of epithelial-mesenchymal transition (EMT) and PI3K/Akt/mTOR signal pathway-related proteins were detected using Western blotting.@*RESULTS@#Online database analysis showed that the expression of P4HA2 was significantly higher in HCC tissues than in normal liver tissues (P < 0.05). The expression levels of P4HA2 mRNA and protein were also significantly higher in HCC cell lines than in normal hepatocytes (P < 0.01). Lentivirus-mediated RNA interference of P4HA2 significantly lowered the expression levels of P4HA2 mRNA and protein in the hepatoma cells (P < 0.05) and caused obvious inhibition of cell proliferation, migration and invasion. P4HA2 knockdown significantly increased the expression of E-cadherin protein, lowered the expressions of N-cadherin and Snail, and obviously decreased the expressions of phosphorylated PI3K, AKT and mTOR (P < 0.05).@*CONCLUSION@#P4HA2 enhances the proliferation, migration, invasion, and EMT of hepatoma cells by activating the PI3K/Akt/mTOR signaling pathway to promote the occurrence and progression of liver cancer.


Subject(s)
Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Humans , Liver Neoplasms/pathology , Phosphatidylinositol 3-Kinases/metabolism , Prolyl Hydroxylases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , RNA, Messenger , Signal Transduction , TOR Serine-Threonine Kinases/metabolism
7.
Article in Chinese | WPRIM | ID: wpr-936343

ABSTRACT

OBJECTIVE@#To explore the mechanism underlying the hepatoprotective effect of dihydromyricetin (DMY) against lipid accumulation in light of the lipophagy pathway and the inhibitory effect of DMY on HepG2 cell proliferation.@*METHODS@#LO2 cells were cultured in the presence of 10% FBS for 24 h and treated with 100 μg/mL DMY, or exposed to 50% FBS for 24 h followed by treatment with 50, 100, or 200 μg/mL DMY; the cells in recovery group were cultured in 50% FBS for 24 h and then in 10% FBS for another 24 h. Oil red O staining was used to observe the accumulation of lipid droplets in the cells, and the levels of TC, TG, and LDL and activities of AST, ALT and LDH were measured. The expression of LC3 protein was detected using Western blotting. AO staining and transmission electron microscopy were used to determine the numbers of autophagolysosomes and autophagosomes, respectively. The formation of autophagosomes was observed with MDC staining, and the mRNA expression levels of LC3, ATG7, AMPK, mTOR, p62 and Beclin1 were determined with q-PCR. Flow cytometry was performed to analyze the effect of 50, 100, and 200 μg/mL DMY on cell cycle and apoptosis of HepG2 cells; DNA integrity in the treated cells was examined with cell DNA fragmentation test.@*RESULTS@#DMY treatment and pretreatment obviously inhibited lipid accumulation and reduced the levels of TC, TG, LDL and enzyme activities of AST, ALT and LDH in LO2 cells (P < 0.05). In routinely cultured LO2 cells, DMY significantly promoted the formation of autophagosomes and autophagolysosomes and upregulated the expression of LC3 protein. DMY obviously attenuated high FBS-induced inhibition of autophagosome formation in LO2 cells, up- regulated the mRNA levels of LC3, ATG7, Beclin1 and AMPK, and downregulated p62 and mTOR mRNA levels (P < 0.05 or 0.01). In HepG2 cells, DMY caused obvious cell cycle arrest, inhibited cell proliferation, and induced late apoptosis and DNA fragmentation.@*CONCLUSION@#DMY reduces lipid accumulation in LO2 cells by regulating the AMPK/ mTOR-mediated lipophagy pathway and inhibits the proliferation of HepG2 by causing cell cycle arrest and promoting apoptosis.


Subject(s)
AMP-Activated Protein Kinases/metabolism , Autophagy , Beclin-1 , Cell Proliferation , Flavonols , Hep G2 Cells , Humans , Lipids , RNA, Messenger , Signal Transduction , TOR Serine-Threonine Kinases/metabolism
8.
Article in Chinese | WPRIM | ID: wpr-936337

ABSTRACT

RNA binding protein (RBP) plays a key role in gene regulation and participate in RNA translation, modification, splicing, transport and other important biological processes. Studies have shown that abnormal expression of RBP is associated with a variety of diseases. The Musashi (Msi) family of mammals is an evolutionarily conserved and powerful RBP, whose members Msi1 and Msi2 play important roles in the regulation of stem cell activity and tumor development. The Msi family members regulate a variety of biological processes by binding and regulating mRNA translation, stability and downstream cell signaling pathways, and among them, Msi2 is closely related to embryonic growth and development, maintenance of tumor stem cells and development of hematological tumors. Accumulating evidence has shown that Msi2 also plays a crucial role in the development of solid tumors, mainly by affecting the proliferation, invasion, metastasis and drug resistance of tumors, involving Wnt/β-catenin, TGF-β/SMAD3, Akt/mTOR, JAK/STAT, Numb and their related signaling pathways (Notch, p53, and Hedgehog pathway). Preclinical studies of Msi2 gene as a therapeutic target for tumor have achieved preliminary results. This review summarizes the molecular structure, physiological function, role of Msi2 in the development and progression of various solid tumors and the signaling pathways involved.


Subject(s)
Animals , Hedgehog Proteins , Mammals/metabolism , Neoplasms/genetics , Neoplastic Stem Cells , RNA-Binding Proteins/metabolism , Signal Transduction
9.
Article in Chinese | WPRIM | ID: wpr-936334

ABSTRACT

OBJECTIVE@#To explore the mechanism by which estradiol modulates the immunophenotype of macrophages through the endoplasmic reticulum stress pathway.@*METHODS@#Peritoneal macrophages isolated from C57 mice were cultured in the presence of 60 ng/mL interferon-γ (IFN-γ) followed by treatment with estradiol (1.0 nmol/L) alone, estradiol with estrogen receptor antagonist (Acolbifene, 4 nmol/L), estradiol with IRE1α inhibitor (4 μ 8 C), or estradiol with IRE1α agonist. After the treatments, the expression levels of MHC-Ⅱ, iNOS and endoplasmic reticulum stress marker proteins IRE1α, eIF2α and ATF6 in the macrophages were detected with Western blotting, and the mRNA levels of TGF-β, IL-6, IL-10 and TNF-α were detected with RT-PCR.@*RESULTS@#Estrogen treatment of the macrophages significantly decreased the expressions of M1-related proteins MHC-Ⅱ (P=0.021) and iNOS (P < 0.001) and the mRNA expressions of TNF-α (P=0.003) and IL-6 (P=0.004), increased the mRNA expression of TGF-β (P=0.002) and IL-10 (P=0.008), and up-regulated the protein expressions of IRE1α (P < 0.001) and its downstream transcription factor XBP-1 (P < 0.001). Addition of the estrogen inhibitor obviously blocked the effect of estrogen. Compared with estrogen treatment alone, combined treatment of the macrophages with estrogen and the IRE1α inhibitor 4 μ 8 C significantly up-regulated the protein expressions of MHC-Ⅱ (P=0.002) and iNOS (P=0.003) and the mRNA expressions of TNF-α (P=0.003) and IL-6 (P=0.024), and obviously down-regulated the mRNA expression of TGF-β (P < 0.001) and IL-10 (P < 0.001); these changes were not observed in cells treated with estrogen and the IRE1α agonist.@*CONCLUSION@#Estrogen can inhibit the differentiation of murine macrophages into a pro-inflammatory phenotype by up-regulating the IRE1α-XBP-1 signaling axis, thereby producing an inhibitory effect on inflammatory response.


Subject(s)
Animals , Cell Differentiation/drug effects , Endoribonucleases/metabolism , Estradiol/pharmacology , Estrogens/metabolism , Interleukin-10 , Interleukin-6/metabolism , Macrophages, Peritoneal/metabolism , Mice , Phenotype , Protein Serine-Threonine Kinases/metabolism , RNA, Messenger/metabolism , Signal Transduction/drug effects , Transforming Growth Factor beta/metabolism , Tumor Necrosis Factor-alpha/metabolism , Up-Regulation/drug effects , X-Box Binding Protein 1/metabolism
10.
Article in Chinese | WPRIM | ID: wpr-936325

ABSTRACT

OBJECTIVE@#To investigate the effect of triptolide (TPL) on inflammatory response and migration of fibroblast like synovial cells (FLS) in rheumatoid arthritis (RA-FLS) and the mechanism of circular noncoding RNA (circRNA) 0003353 for mediating this effect.@*METHODS@#We collected peripheral blood mononuclear cells (PBMCs) and serum samples from 50 hospitalized RA patients and 30 healthy individuals for detecting the expression of circRNA 0003353, immune and inflammatory indexes (ESR, CRP, RF, anti-CCP, IgA, IgG, IgM, C3, and C4) and DAS28 score. Cultured RA-FLS was treated with 10 ng/mL TPL and transfected with a circRNA 0003353 overexpression plasmid, and cell counting kit-8 (CCK-8) assay and Transwell assay were used to detect the changes in the viability and migration of the cells. Enzyme-linked immunosorbent assay (ELISA) was used to examine the cytokines IL-4, IL-6, and IL-17, and real-time fluorescence quantitative PCR (RT-qPCR) was performed to detect the expression of circRNA 003353; Western blotting was used to detect the expressions of p-JAK2, pSTAT3, JAK2 and STAT3 proteins in the treated cells.@*RESULTS@#The expression of circRNA 0003353 was significantly increased in PBMCs from RA patients and showed a good performance in assisting the diagnosis of RA (AUC=90.5%, P < 0.001, 95% CI: 0.83-0.98). CircRNA 0003353 expression was positively correlated with ESR, RF and DAS28 (P < 0.05). Treatment with TPL significantly decreased the expression of circRNA 0003353, suppressed the viability and migration ability, decreased the expressions of IL-6 and IL-17, and increased the expression IL-4 in cultured RA-FLS in a time-dependent manner (P < 0.01). TNF-α stimulation of RA-FLS significantly increased the ratios of p-JAK2/JAK2 and p-STAT3/STAT3, which were obviously lowered by TPL treatment (P < 0.01). TPL-treated RA-FLS overexpressing circRNA 0003353 showed significantly increased cell viability and migration ability with decreased IL-4 expression and increased IL-6 and IL-17 expressions and ratios of p-JAK2/ JAK2 and p-STAT3/STAT3 (P < 0.01).@*CONCLUSION@#The expression of circRNA 0003353 is increased in PBMCs in RA patients and in RA-FLS. TPL treatment can regulate JAK2/STAT3 signal pathway and inhibit the inflammatory response and migration of RA-FLS through circRNA 0003353.


Subject(s)
Arthritis, Rheumatoid/pathology , Cells, Cultured , Diterpenes/pharmacology , Epoxy Compounds/pharmacology , Fibroblasts/pathology , Humans , Interleukin-17/metabolism , Interleukin-4/metabolism , Interleukin-6/metabolism , Janus Kinase 2/metabolism , Leukocytes, Mononuclear/metabolism , Phenanthrenes/pharmacology , RNA, Circular/metabolism , STAT3 Transcription Factor/metabolism , Signal Transduction/drug effects , Synovial Membrane/pathology
11.
Article in Chinese | WPRIM | ID: wpr-936312

ABSTRACT

OBJECTIVE@#To investigate the changes in autophagy of mesenchymal stem cells (MSCs) from patients with ankylosing spondylitis and explore the mechanism for decreased autophagy in ASMSCs.@*METHODS@#MSCs collected from 14 patients with AS (ASMSCs) and from 15 healthy donors (HDMSCs) were cultured in the absence or presence of 25 ng/mL TNF-α for 6 h. Autophagy of the cells was determined by immunofluorescence staining of GFP-LC3B, and the results were confirmed by detecting the protein expressions of autophagy markers LC3 II/LC3 I and P62. The mRNA expressions of the related genes were detected using qRT-PCR, and the protein expressions of the autophagy markers and signaling pathway-related molecules were determined with Western blotting. TG100713 was used to block the PI3K/AKT/mTOR signal pathway, and its effect on autophagy of ASMSCs was evaluated.@*RESULTS@#ASMSCs showed significantly weaker GFP-LC3B puncta staining and lower protein expression levels of LC3 II/LC3 I but higher levels of P62 protein (P < 0.05), indicating a decreased autophagy capacity as compared with HDMSCs. TNF-α-induced ASMSCs showed significantly higher protein expressions of p-PI3K/ PI3K, p-AKT/AKT and p-mTOR/mTOR than HDMSCs (P < 0.05), suggesting hyperactivation of the PI3K/AKT/mTOR signaling pathway in ASMSCs. Blocking PI3K/AKT/mTOR signaling with TG100713 eliminated the difference in TNF-α-induced autophagy between HDMSCs and ASMSCs.@*CONCLUSION@#In patients with AS, hyperactivation of the PI3K/AKT/mTOR signaling pathway results in decreased autophagy of the MSCs and potentially contributes to chronic inflammation.


Subject(s)
Autophagy , Humans , Mesenchymal Stem Cells/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction , Spondylitis, Ankylosing , TOR Serine-Threonine Kinases/metabolism , Tumor Necrosis Factor-alpha/metabolism
12.
Article in Chinese | WPRIM | ID: wpr-936285

ABSTRACT

OBJECTIVE@#To investigate the inhibitory effect of 27-P-coumayl-ursolic acid (27-P-CAUA), the active ingredient in triterpenoids from the leaves of Ilex latifolia Thunb, against breast cancer cells and explore the underlying mechanism.@*METHODS@#CCK-8 assay was used to assess the changes in viability of breast cancer HCC-1806 cells after 27-P-CAUA treatment for 24, 48, or 72 h. The inhibitory effect of 27-P-CAUA on proliferation of the cells was determined by clonogenic assay. JC-1 was used to detect the changes in mitochondrial membrane potential and flow cytometry was performed for analyzing cell apoptosis following 27-P-CAUA treatment. Immunofluorescence assay was used to observe the expression of cl-caspase-3 and P62 in the treated cells. Western blotting was performed to observe the effect of 27-P-CAUA and chloroquine pretreatment on the expressions of LC3I/II, P62 and HER2 signaling pathway proteins in the cells.@*RESULTS@#The results of CCK-8 and clonogenic assays showed that 27-P-CAUA treatment significantly inhibited the proliferation of HCC-1806 cells (P < 0.01) with IC50 values of 81.473, 48.392 and 18.467 μmol/L at 24, 48, and 72 h, respectively. 27-P-CAUA treatment also caused obvious changes in mitochondrial membrane potential (P < 0.01) and induced cell apoptosis in HCC-1806 cells with a 3.34% increase of the early apoptosis rate. Immunofluorescence assay revealed a significant increase of cl-caspase3 expression in 27-P-CAUA-treated HCC-1806 cells, and treatment with 40 μmol/L 27-P-CAUA resulted in significant cell apoptosis (P < 0.01). 27-P-CAUA obviously reduced the expression of LC3II, caused P62 degradation and induced autophagy in HCC-1806 cells. Chloroquine pretreatment obviously blocked the autophagy-inducing effect of 27-P-CAUA. 27-P-CAUA treatment also inhibited the phosphorylation of HER2 and AKT proteins and progressively lowered the expressions of HER2 and phosphorylated AKT protein in HCC-1806 cells (P < 0.01).@*CONCLUSION@#27-P-CAUA can inhibit the proliferation and induce mitochondrial autophagy and apoptosis of HCC-1806 cells by inhibiting the HER2/PI3K/AKT signaling pathway.


Subject(s)
Apoptosis , Autophagy , Breast Neoplasms , Carcinoma, Hepatocellular/metabolism , Cell Line, Tumor , Cell Proliferation , Female , Humans , Liver Neoplasms/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction
13.
Article in English | WPRIM | ID: wpr-929264

ABSTRACT

Abelmoschus manihot (L.) Medik. (A. manihot) is a traditional Chinese herbal medicine with a variety of pharmacological properties. It was first recorded in Jiayou Materia Medica dating back to the Song dynasty to eliminate urinary tract irritation by clearing away heat and diuretic effect. However, its pharmacological action on urinary tract infections has not been investigated. The present study aims to evaluate the anti-inflammatory activity of A. manihot on a mouse model of lipopolysaccharide (LPS)-induced cystitis. The results showed that A. manihot decreased white blood cell (WBC) count in urine sediments of the cystitis mice, alleviated bladder congestion, edema, as well as histopathological damage, reduced the expression levels of tumor necrosis factor-α, interleukin-6, and interleukin-1β simultaneously. Moreover, A. manihot administration significantly downregulated the expression levels of TLR4, MYD88, IκBα, p-IκBα, NF-κB p65, and p-NF-κB p65 in LPS-induced cystitis mice. These findings demonstrated the protective effect of A. manihot against LPS-induced cystitis, which is attributed to its anti-inflammatory profile by suppressing TLR4/MYD88/NF-κB pathways. Our results suggest that A. manihot could be a potential candidate for cystitis treatment.


Subject(s)
Abelmoschus/metabolism , Animals , Anti-Inflammatory Agents/therapeutic use , Cystitis , Female , Humans , Lipopolysaccharides/pharmacology , Male , Mice , Myeloid Differentiation Factor 88/metabolism , NF-KappaB Inhibitor alpha/metabolism , NF-kappa B/metabolism , Signal Transduction , Toll-Like Receptor 4/metabolism
14.
Article in English | WPRIM | ID: wpr-929250

ABSTRACT

To explore the effectiveness and safety of a Chinese medicinal decoction Wuwei Xiaodu Drink (WWXDD) in inhibiting chronic osteomyelitis via regulatory T cells signaling. The effective constitutes of WWXDD and osteomyelitis related genes were screened. Target proteins were cross-validated using the Venny database. GO function and KEGG pathway analysis were performed for target proteins, while pharmacological network was constructed. The bone properties were analyzed by HE staining and the concentrations of immune factors were measured by ELISA. The expression of CTLA-4 and Foxp3 mRNA and STAT5, p-STAT5, CTLA-4 and Foxp3 protein were detected using Real-time PCR and Western blot, respectively. FACS was used to analyze the percentages of cells. A total of 117 genes overlapped between 785 target genes of the active compounds of WWXDD and 912 osteomyelitis related genes. Inflammation-related genes, including IL-6, TNFα, IL-1β and IL-2 showed high connection degree in the drug-compound-disease-target network. GO function and KEGG pathway analysis revealed that 117 intersection genes mainly enriched in virus infection related pathways, immune related pathways and chemokine signaling pathway. Furthermore, the development of chronic osteomyelitis was suppressed in model rats after treatment with WWXDD. Meanwhile, the concentrations of IL-2 and CD4+CD25+Foxp3 Treg percentages together with the levels of p-STAT5, CTLA-4 and Foxp3 were also down-regulated. Furthermore, IL-2 and WWXDD drug-containing serum exhibited opposite effects on regulating IL-2, IL-10, TGF-β1, Foxp3, CTLA4 and STAT5. In addition, a STAT5 phosphorylation inhibitor suppressed the expression of Foxp3 and CTLA-4. WWXDD can treat chronic osteomyelitis through suppressing the main regulating factors of Tregs and interfere its immunodepression. Our results bring a new solution for chronic osteomyelitis.


Subject(s)
Animals , Forkhead Transcription Factors/metabolism , Interleukin-2/metabolism , Osteomyelitis/metabolism , Rats , STAT5 Transcription Factor/metabolism , Signal Transduction , T-Lymphocytes, Regulatory
15.
Journal of Integrative Medicine ; (12): 163-172, 2022.
Article in English | WPRIM | ID: wpr-929221

ABSTRACT

OBJECTIVE@#Moxibustion, a common therapy in traditional Chinese medicine, has potential benefits for treating decreased ovarian reserve (DOR). The present study investigates the protective effect of moxibustion in a rat model of DOR and explores the possible mechanisms.@*METHODS@#Sixty-four female Sprague-Dawley rats were randomly divided into four groups: control, DOR, moxibustion (MOX), and hormone replacement therapy (HRT). The DOR rat model was established by intragastric administration of 50 mg/kg Tripterygium glycoside suspension (TGS), once daily for 14 days. MOX and HRT treatments were given from the day TGS administration was initiated. The ovarian reserve function was evaluated by monitoring the estrus cycle, morphological changes in ovaries, levels of serum estradiol (E2), follicle-stimulating hormone (FSH), luteinizing hormone (LH), and anti-Mullerian hormone (AMH), pregnancy rate and embryo numbers. Terminal-deoxynucleotidyl transferase-mediated nick-end-labeling staining was used to identify ovarian granulosa cell apoptosis, while the protein and mRNA expressions of Bax, B-cell lymphoma-2 (Bcl-2), phosphatidylinositol 3-kinase (PI3K) and protein kinase B (AKT) in ovarian tissues were examined by immunohistochemistry, Western blot and quantitative reverse transcription-polymerase chain reaction.@*RESULTS@#Compared with the DOR group, MOX improved the disordered estrous cycle, promoted follicular growth, reduced the number of atresia follicles, increased the concentrations of serum E2 and AMH, and decreased serum FSH and LH concentrations. More importantly, the pregnancy rate and embryo numbers in DOR rats were both upregulated in the MOX treatment group, compared to the untreated DOR model. Further, we found that the MOX group had reduced apoptosis of ovarian granulosa cells, increased Bcl-2 expression and reduced expression of Bax. Furthermore, the PI3K/AKT signaling pathway was triggered by the moxibustion treatment.@*CONCLUSION@#Moxibustion improved ovarian function and suppressed apoptosis of ovarian granulosa cells in a rat model of DOR induced by TGS, and the mechanism may involve the PI3K/AKT signaling pathway.


Subject(s)
Animals , Female , Follicle Stimulating Hormone , Luteinizing Hormone , Moxibustion , Ovarian Reserve , Phosphatidylinositol 3-Kinase/pharmacology , Phosphatidylinositol 3-Kinases/metabolism , Pregnancy , Proto-Oncogene Proteins c-akt/pharmacology , Rats , Rats, Sprague-Dawley , Signal Transduction , bcl-2-Associated X Protein/genetics
16.
Protein & Cell ; (12): 90-101, 2022.
Article in English | WPRIM | ID: wpr-929171

ABSTRACT

The cyclic GMP-AMP (cGAMP) synthase (cGAS) has been identified as a cytosolic double stranded DNA sensor that plays a pivotal role in the type I interferon and inflammation responses via the STING-dependent signaling pathway. In the past several years, a growing body of evidence has revealed that cGAS is also localized in the nucleus where it is associated with distinct nuclear substructures such as nucleosomes, DNA replication forks, the double-stranded breaks, and centromeres, suggesting that cGAS may have other functions in addition to its role in DNA sensing. However, while the innate immune function of cGAS is well established, the non-canonical nuclear function of cGAS remains poorly understood. Here, we review our current understanding of the complex nature of nuclear cGAS and point to open questions on the novel roles and the mechanisms of action of this protein as a key regulator of cell nuclear function, beyond its well-established role in dsDNA sensing and innate immune response.


Subject(s)
Cell Nucleus/immunology , Humans , Immunity, Innate , Nucleotidyltransferases/immunology , Signal Transduction/immunology
17.
Protein & Cell ; (12): 6-25, 2022.
Article in English | WPRIM | ID: wpr-929158

ABSTRACT

The serine/threonine p21-activated kinases (PAKs), as main effectors of the Rho GTPases Cdc42 and Rac, represent a group of important molecular switches linking the complex cytoskeletal networks to broad neural activity. PAKs show wide expression in the brain, but they differ in specific cell types, brain regions, and developmental stages. PAKs play an essential and differential role in controlling neural cytoskeletal remodeling and are related to the development and fate of neurons as well as the structural and functional plasticity of dendritic spines. PAK-mediated actin signaling and interacting functional networks represent a common pathway frequently affected in multiple neurodevelopmental and neurodegenerative disorders. Considering specific small-molecule agonists and inhibitors for PAKs have been developed in cancer treatment, comprehensive knowledge about the role of PAKs in neural cytoskeletal remodeling will promote our understanding of the complex mechanisms underlying neurological diseases, which may also represent potential therapeutic targets of these diseases.


Subject(s)
Animals , Cytoskeleton/genetics , Humans , Nervous System Diseases/genetics , Neurons/enzymology , Signal Transduction , p21-Activated Kinases/metabolism
18.
Article in English | WPRIM | ID: wpr-929149

ABSTRACT

Sphingosine-1-phosphate (S1P) is an important lipid mediator that regulates a diverse range of intracellular cell signaling pathways that are relevant to tissue engineering and regenerative medicine. However, the precise function of S1P in dental pulp stem cells (DPSCs) and its osteogenic differentiation remains unclear. We here investigated the function of S1P/S1P receptor (S1PR)-mediated cellular signaling in the osteogenic differentiation of DPSCs and clarified the fundamental signaling pathway. Our results showed that S1P-treated DPSCs exhibited a low rate of differentiation toward the osteogenic phenotype in association with a marked reduction in osteogenesis-related gene expression and AKT activation. Of note, both S1PR1/S1PR3 and S1PR2 agonists significantly downregulated the expression of osteogenic genes and suppressed AKT activation, resulting in an attenuated osteogenic capacity of DPSCs. Most importantly, an AKT activator completely abrogated the S1P-mediated downregulation of osteoblastic markers and partially prevented S1P-mediated attenuation effects during osteogenesis. Intriguingly, the pro-inflammatory TNF-α cytokine promoted the infiltration of macrophages toward DPSCs and induced S1P production in both DPSCs and macrophages. Our findings indicate that the elevation of S1P under inflammatory conditions suppresses the osteogenic capacity of the DPSCs responsible for regenerative endodontics.


Subject(s)
Cell Differentiation , Cell Proliferation , Cells, Cultured , Dental Pulp/metabolism , Lysophospholipids , Osteogenesis , Proto-Oncogene Proteins c-akt/metabolism , Signal Transduction , Sphingosine/analogs & derivatives , Stem Cells
19.
Article in English | WPRIM | ID: wpr-929054

ABSTRACT

Marine fungi are important members of the marine microbiome, which have been paid growing attention by scientists in recent years. The secondary metabolites of marine fungi have been reported to contain rich and diverse compounds with novel structures (Chen et al., 2019). Aspergillus terreus, the higher level marine fungus of the Aspergillus genus (family of Trichocomaceae, order of Eurotiales, class of Eurotiomycetes, phylum of Ascomycota), is widely distributed in both sea and land. In our previous study, the coral-derived A. terreus strain C23-3 exhibited potential in producing other biologically active (with antioxidant, acetylcholinesterase inhibition, and anti-inflammatory activity) compounds like arylbutyrolactones, territrems, and isoflavones, and high sensitivity to the chemical regulation of secondary metabolism (Yang et al., 2019, 2020; Nie et al., 2020; Ma et al., 2021). Moreover, we have isolated two different benzaldehydes, including a benzaldehyde with a novel structure, from A. terreus C23-3 which was derived from Pectinia paeonia of Xuwen, Zhanjiang City, Guangdong Province, China.


Subject(s)
Acetylcholinesterase/metabolism , Animals , Anthozoa/microbiology , Anti-Inflammatory Agents/pharmacology , Aspergillus/chemistry , Benzaldehydes/pharmacology , Mice , Signal Transduction
20.
Article in English | WPRIM | ID: wpr-929038

ABSTRACT

Mesenchymal stem/stromal cell (MSC)‍-based therapy has been regarded as one of the most revolutionary breakthroughs in the history of modern medicine owing to its myriad of immunoregulatory and regenerative properties. With the rapid progress in the fields of osteo- and musculoskeletal therapies, the demand for MSC-based treatment modalities is becoming increasingly prominent. In this endeavor, researchers around the world have devised new and innovative techniques to support the proliferation of MSCs while minimizing the loss of hallmark features of stem cells. One such example is electromagnetic field (EMF) exposure, which is an alternative approach with promising potential. In this review, we present a critical discourse on the efficiency, practicability, and limitations of some of the relevant methods, with insurmountable evidence backing the implementation of EMF as a feasible strategy for the clinically relevant expansion of MSCs.


Subject(s)
Cell Differentiation , Cell Proliferation , Electromagnetic Fields , Mesenchymal Stem Cells , Signal Transduction
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