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1.
Protein & Cell ; (12): 788-809, 2021.
Article in English | WPRIM | ID: wpr-922475

ABSTRACT

Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and is the fourth-leading cause of cancer-related deaths worldwide. HCC is refractory to many standard cancer treatments and the prognosis is often poor, highlighting a pressing need to identify biomarkers of aggressiveness and potential targets for future treatments. Kinesin family member 2C (KIF2C) is reported to be highly expressed in several human tumors. Nevertheless, the molecular mechanisms underlying the role of KIF2C in tumor development and progression have not been investigated. In this study, we found that KIF2C expression was significantly upregulated in HCC, and that KIF2C up-regulation was associated with a poor prognosis. Utilizing both gain and loss of function assays, we showed that KIF2C promoted HCC cell proliferation, migration, invasion, and metastasis both in vitro and in vivo. Mechanistically, we identified TBC1D7 as a binding partner of KIF2C, and this interaction disrupts the formation of the TSC complex, resulting in the enhancement of mammalian target of rapamycin complex1 (mTORC1) signal transduction. Additionally, we found that KIF2C is a direct target of the Wnt/β-catenin pathway, and acts as a key factor in mediating the crosstalk between Wnt/β-catenin and mTORC1 signaling. Thus, the results of our study establish a link between Wnt/β-catenin and mTORC1 signaling, which highlights the potential of KIF2C as a therapeutic target for the treatment of HCC.


Subject(s)
Adult , Aged , Animals , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cell Movement , Cell Proliferation , Epithelial-Mesenchymal Transition/genetics , Female , Gene Expression Regulation, Neoplastic , Humans , Intracellular Signaling Peptides and Proteins/metabolism , /metabolism , Liver Neoplasms/pathology , Male , Mice , Mice, Inbred BALB C , Middle Aged , Neoplasm Staging , Prognosis , Protein Binding , RNA, Small Interfering/metabolism , Survival Analysis , Tumor Burden , Wnt Signaling Pathway , Xenograft Model Antitumor Assays , beta Catenin/metabolism
2.
Article in Chinese | WPRIM | ID: wpr-921814

ABSTRACT

To explore the regulatory effects of Xinfeng Capsules(XFC) on the apoptosis of synovial fibroblasts(FLS) and inflammation in rheumatoid arthritis(RA) via lncRNA MAPKAPK5-AS1(MK5-AS1). Thirty healthy people and 30 patients with RA due to spleen deficiency and dampness exuberance were collected for extracting the peripheral blood mononuclear cells(PBMCs) before and after XFC treatment, which were used to observe the correlation between MK5-AS1 and clinical indicators as well as MK5-AS1 expression before and after XFC treatment. Following the establishment of RA-FLS cell line and the preparation of XFC-containing serum, MK5-AS1-overexpression plasmid was constructed and transfected into RA-FLS for investigating the efficacy of XFC-containing serum in regulating inflammation and apoptosis of RA-FLS via MK5-AS1. The expression of MK5-AS1 in PBMCs of patients with RA due to spleen deficiency and dampness exuberance was decreased(P<0.001). The ROC curve analysis revealed the AUC of 83.9%. Correlation analysis showed that MK5-AS1 was negatively correlated with ESR, CRP, RF, CCP, and spleen deficiency and dampness exuberance syndrome score. The expression of MK5-AS1 increased significantly after XFC treatment(P<0.001). As demonstrated by association analysis, XFC decreased MK5-AS1, ESR, CRP, RF, and spleen deficiency and dampness exuberance syndrome score, with the degree of support all greater than 83%, confidence greater than 80%, and lift greater than 1. The results of RT-qPCR showed that the MK5-AS1 RNA expression significantly decreased after TNF-α stimulation(P<0.01), which, however, increased significantly after the intervention with XFC-containing serum(P<0.05). Such expression rose again after the transfection of pcDNA3.1-MK5-AS1(P<0.01). ELISA results showed that TNF-α stimulation elevated the expression of pro-inflammatory factor IL-17 but lowered the expression of anti-inflammatory factor IL-4(P<0.01). After intervention with XFC-containing serum, the expression of IL-17 decreased while that of IL-4 increased(P<0.01). The transfection of pcDNA3.1-MK5-AS1 contributed to the reduction in IL-17 expression but the elevation in IL-4 expression(P<0.01). The immunofluorescence(IF) findings demonstrated that the expression of pro-apoptotic protein Bax was down-regulated, whereas that of the anti-apoptotic protein Bcl-2 was up-regulated after TNF-α stimulation(P<0.01). After the intervention with XFC-containing serum, the Bax expression was increased, while Bcl-2 expression was decreased(P<0.01), which were remarkably collaborated by the transfection of pcDNA3.1-MK5-AS1(P<0.05). The expression of MK5-AS1 is significantly decreased in both RA-PBMCs and RA-FLS, implying that XFC inhibits inflammatory reaction and promotes the apoptosis in RA by regulating the expression of MK5-AS1.


Subject(s)
Apoptosis , Arthritis, Rheumatoid/genetics , Capsules , Drugs, Chinese Herbal , Fibroblasts , Humans , Inflammation/genetics , Intracellular Signaling Peptides and Proteins , Leukocytes, Mononuclear , RNA, Long Noncoding/genetics
3.
Chinese Medical Journal ; (24): 2340-2352, 2021.
Article in English | WPRIM | ID: wpr-921125

ABSTRACT

BACKGROUND@#Emerging evidence indicates that the sineoculis homeobox homolog 1-eyes absent homolog 1 (SIX1-EYA1) transcriptional complex significantly contributes to the pathogenesis of multiple cancers by mediating the expression of genes involved in different biological processes, such as cell-cycle progression and metastasis. However, the roles of the SIX1-EYA1 transcriptional complex and its targets in colorectal cancer (CRC) are still being investigated. This study aimed to investigate the roles of SIX1-EYA1 in the pathogenesis of CRC, to screen inhibitors disrupting the SIX1-EYA1 interaction and to evaluate the efficiency of small molecules in the inhibition of CRC cell growth.@*METHODS@#Real-time quantitative polymerase chain reaction and western blotting were performed to examine gene and protein levels in CRC cells and clinical tissues (collected from CRC patients who underwent surgery in the Department of Integrated Traditional and Western Medicine, West China Hospital of Sichuan University, between 2016 and 2018, n = 24). In vivo immunoprecipitation and in vitro pulldown assays were carried out to determine SIX1-EYA1 interaction. Cell proliferation, cell survival, and cell invasion were determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, clonogenic assay, and Boyden chamber assay, respectively. The Amplified Luminescent Proximity Homogeneous Assay Screen (AlphaScreen) method was used to obtain small molecules that specifically disrupted SIX1-EYA1 interaction. CRC cells harboring different levels of SIX1/EYA1 were injected into nude mice to establish tumor xenografts, and small molecules were also injected into mice to evaluate their efficiency to inhibit tumor growth.@*RESULTS@#Both SIX1 and EYA1 were overexpressed in CRC cancerous tissues (for SIX1, 7.47 ± 3.54 vs.1.88 ± 0.35, t = 4.92, P = 0.008; for EYA1, 7.61 ± 2.03 vs. 2.22 ± 0.45, t = 6.73, P = 0.005). The SIX1/EYA1 complex could mediate the expression of two important genes including cyclin A1 (CCNA1) and transforming growth factor beta 1 (TGFB1) by binding to the myocyte enhancer factor 3 consensus. Knockdown of both SIX1 and EYA1 could decrease cell proliferation, cell invasion, tumor growth, and in vivo tumor growth (all P < 0.01). Two small molecules, NSC0191 and NSC0933, were obtained using AlphaScreen and they could significantly inhibit the SIX1-EYA1 interaction with a half-maximal inhibitory concentration (IC50) of 12.60 ± 1.15 μmol/L and 83.43 ± 7.24 μmol/L, respectively. Administration of these two compounds could significantly repress the expression of CCNA1 and TGFB1 and inhibit the growth of CRC cells in vitro and in vivo.@*CONCLUSIONS@#Overexpression of the SIX1/EYA1 complex transactivated the expression of CCNA1 and TGFB1, causing the pathogenesis of CRC. Pharmacological inhibition of the SIX1-EYA1 interaction with NSC0191 and NSC0933 significantly inhibited CRC cell growth by affecting cell-cycle progression and metastasis.


Subject(s)
Animals , Cell Line, Tumor , Cell Proliferation , Colorectal Neoplasms/genetics , Gene Expression Regulation, Neoplastic , Genes, Homeobox , Homeodomain Proteins/metabolism , Humans , Intracellular Signaling Peptides and Proteins , Mice , Mice, Nude , Nuclear Proteins/genetics , Protein Tyrosine Phosphatases/genetics
4.
Article in English | WPRIM | ID: wpr-878450

ABSTRACT

OBJECTIVES@#This study aims to investigate the effect of the regulator of G-protein signaling 2 (RGS2) on the proliferation and invasion of oral squamous cell carcinoma (OSCC) cells and its potential molecular mechanism. Metho⁃ds The expression status and clinical significance of RGS2 in head and neck squamous cell carcinomas and matched adjacent normal tissues were evaluated using TCGA database. Three OSCC cell lines (i.e., SCC-9, Cal27, and Fadu) were overexpressed with RGS2, and the effect of RGS2 on cell proliferation and invasion was determined using the Transwell, clone formation, and cell counting kit (CCK)-8 assays. Moreover, the yeast two-hybrid scree-ning and co-immunoprecipitation (Co-IP) assays were conducted to detect the correlation of RGS2, four and a half LIM domains protein 1 (FHL1), and damage DNA-binding protein 1 (DDB1).@*RESULTS@#The expression level of RGS2 in OSCC was significantly lower than that in matched adjacent normal tissues (@*CONCLUSIONS@#RGS2 plays an important role in the inhibition of OSCC proliferation and invasion. The structure stability of RGS2 is competitively regulated by FHL1 and DDB1.


Subject(s)
Carcinoma, Squamous Cell , Cell Line, Tumor , Cell Movement , Cell Proliferation , GTP-Binding Proteins , Head and Neck Neoplasms , Humans , Intracellular Signaling Peptides and Proteins , LIM Domain Proteins , Mouth Neoplasms , Muscle Proteins , Squamous Cell Carcinoma of Head and Neck
5.
Säo Paulo med. j ; 138(2): 133-139, Mar.-Apr. 2020. tab, graf
Article in English | SES-SP, LILACS, SES-SP | ID: biblio-1139677

ABSTRACT

ABSTRACT BACKGROUND: Diabetic nephropathy is a common complication of chronic kidney disease (CKD). ­Inflammation in the kidneys is crucial for promoting development and progression of this complication. Wnt member 5a (Wnt5a) and secreted frizzled-related protein 5 (Sfrp5) are proinflammatory proteins associated with insulin resistance and chronic low-grade adipose tissue inflammation. OBJECTIVE: To determine the correlation between serum Sfrp5 and Wnt5a concentrations and glomerular filtration rate in patients with type 2 diabetes mellitus and CKD. DESIGN AND SETTING: Cross-sectional, comparative and observational study in the Department of Endocrinology, Civil Hospital, Culiacán, Sinaloa, Mexico. METHODS: Eighty individuals with chronic kidney disease were recruited. Their serum Sfrp5 and Wnt5a concentrations were quantified using the enzyme-linked immunosorbent assay (ELISA) test. The statistical analysis consisted of the Mann-Whitney U test for independent samples and Spearman correlation, with statistical significance of P < 0.05. RESULTS: Serum Sfrp5 concentration continually increased through the stages of CKD progression, whereas serum Wnt5a concentration presented its highest levels in stage 3 CKD. Negative correlations between estimated glomerular filtration rate (eGFR) and serum concentrations of Sfrp5 (r = -0434, P = 0.001) and Wnt5a (r = -0481, P = 0.001) were found. CONCLUSIONS: There were negative correlations between serum Sfrp5 and Wnt5a concentrations and eGFR at each stage of CKD, with higher levels in female patients. This phenomenon suggests that Sfrp5 and Wnt5a might be involved in development and evolution towards end-stage renal disease.


Subject(s)
Humans , Female , Intracellular Signaling Peptides and Proteins/blood , Diabetes Mellitus, Type 2 , Renal Insufficiency, Chronic , Kidney Failure, Chronic , Cross-Sectional Studies , Eye Proteins , Glomerular Filtration Rate , Membrane Proteins , Mexico
6.
Arq. neuropsiquiatr ; 78(1): 21-27, Jan. 2020. graf
Article in English | LILACS | ID: biblio-1088987

ABSTRACT

Abstract Objective: The phytohormone abscisic acid (ABA) as a signaling molecule exists in various types of organisms from early multicellular to animal cells and tissues. It has been demonstrated that ABA has an antinociceptive effect in rodents. The present study was designed to assess the possible role of PKA and phosphorylated ERK (p-ERK) on the antinociceptive effects of intrathecal (i.t.) ABA in male Wistar rats. Methods: The animals were cannulated intrathecally and divided into different experimental groups (n=6‒7): Control (no surgery), vehicle (received ABA vehicle), ABA-treated groups (received ABA in doses of 10 or 20 µg/rat), ABA plus H.89 (PKA inhibitor)-treated group which received the inhibitor 15 min prior to the ABA injection. Tail-flick and hot-plate tests were used as acute nociceptive stimulators to assess ABA analgesic effects. p-ERK was evaluated in the dorsal portion of the spinal cord using immunoblotting. Results: Data showed that a microinjection of ABA (10 and 20 µg/rat, i.t.) significantly increased the nociceptive threshold in tail flick and hot plate tests. The application of PKA inhibitor (H.89, 100 nM/rat) significantly inhibited ABA-induced analgesic effects. Expression of p-ERK was significantly decreased in ABA-injected animals, which were not observed in the ABA+H.89-treated group. Conclusions: Overall, i.t. administration of ABA (10 µg/rat) induced analgesia and p-ERK down-expression likely by involving the PKA-dependent mechanism.


Resumo Objetivo: O ácido fito-hormônio abscísico (ABA) existe como molécula sinalizadora em vários tipos de organismos, de multicelulares a células e tecidos animais. Foi demonstrado que o ABA tem efeito antinociceptivo em roedores. O presente estudo foi desenhado para avaliar o possível papel da PKA e da ERK fosforilada (p-ERK) nos efeitos antinociceptivos do ABA intratecal (i.t.) em ratos Wistar machos. Métodos: Os animais foram canulados por via i.t. e divididos em diferentes grupos experimentais (n=6‒7): controle (sem cirurgia), veículo (veículo ABA recebido), grupos tratados com ABA (recebeu ABA em doses de 10 ou 20 µg/rato), grupo tratado com ABA mais H.89 (inibidor de PKA) que recebeu o inibidor 15 minutos antes da injeção de ABA. Os testes de movimento da cauda e placa quente foram utilizados como estimuladores nociceptivos agudos para avaliar os efeitos analgésicos da ABA. A p-ERK foi avaliada na porção dorsal da medula espinhal por imunotransferência. Resultados: A microinjeção de ABA (10 e 20 µg/rato, i.t.) aumentou significativamente o limiar nociceptivo nos testes de movimento da cauda e placa quente. A aplicação de inibidor de PKA (H.89, 100 nM/rato) inibiu significativamente os efeitos analgésicos induzidos por ABA. A expressão de p-ERK diminuiu significativamente em animais injetados com ABA que não foram observados no grupo tratado com ABA+H.89. Conclusões: No geral, a administração i.t. de ABA (10 µg/rato) induziu a analgesia e expressão negativa de p-ERK provavelmente envolvendo mecanismo dependente de PKA.


Subject(s)
Animals , Male , Plant Growth Regulators/pharmacology , Spinal Cord/metabolism , Abscisic Acid/pharmacology , Cyclic AMP-Dependent Protein Kinases/drug effects , Extracellular Signal-Regulated MAP Kinases/drug effects , Analgesics/pharmacology , Reference Values , Spinal Cord/drug effects , Time Factors , Blotting, Western , Reproducibility of Results , Rats, Wistar , Cyclic AMP-Dependent Protein Kinases/analysis , Extracellular Signal-Regulated MAP Kinases/analysis , Intracellular Signaling Peptides and Proteins/pharmacology
7.
Article in Chinese | WPRIM | ID: wpr-828927

ABSTRACT

Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial pneumonia characterized by progressive accumulation of fibroblastic foci and destruction of the alveolar structure. Due to an incomplete understanding of the mechanism of the occurrence and progression of IPF, currently no effective means have been available for its early screening or treatment. With a poor overall prognosis, the patients with IPF have a median survival of only 2-4 years. In recent years, several studies have confirmed that dozens of molecules are involved in the development of IPF and can be used as potential biomarkers. These biomarkers play important roles in early diagnosis (such as SP-D, MMP-7, and osteopontin), prognostic evaluation (such as telomerase length, KL-6, mtDNA, HSP-70, LOXL2, CXCL13, miRNA, ICAM-1, and CCL18), and guiding treatment of IPF (such as TOLLIP rs3750920 genotype, SAMS score, and SP-D), and also provide potential therapeutic targets (such as TERT, TERR, RTEC, and PARN).


Subject(s)
Amino Acid Oxidoreductases , Biomarkers , Disease Progression , Humans , Idiopathic Pulmonary Fibrosis , Intracellular Signaling Peptides and Proteins , Matrix Metalloproteinase 7 , Prognosis
8.
Article in Chinese | WPRIM | ID: wpr-828508

ABSTRACT

Idiopathic pulmonary fibrosis (IPF) is a chronic interstitial pneumonia characterized by progressive accumulation of fibroblastic foci and destruction of the alveolar structure. Due to an incomplete understanding of the mechanism of the occurrence and progression of IPF, currently no effective means have been available for its early screening or treatment. With a poor overall prognosis, the patients with IPF have a median survival of only 2-4 years. In recent years, several studies have confirmed that dozens of molecules are involved in the development of IPF and can be used as potential biomarkers. These biomarkers play important roles in early diagnosis (such as SP-D, MMP-7, and osteopontin), prognostic evaluation (such as telomerase length, KL-6, mtDNA, HSP-70, LOXL2, CXCL13, miRNA, ICAM-1, and CCL18), and guiding treatment of IPF (such as TOLLIP rs3750920 genotype, SAMS score, and SP-D), and also provide potential therapeutic targets (such as TERT, TERR, RTEC, and PARN).


Subject(s)
Amino Acid Oxidoreductases , Metabolism , Biomarkers , Disease Progression , Humans , Idiopathic Pulmonary Fibrosis , Diagnosis , Therapeutics , Intracellular Signaling Peptides and Proteins , Metabolism , Prognosis
9.
Article in English | WPRIM | ID: wpr-827414

ABSTRACT

OBJECTIVES@#To observe the electrophysiological changes of astrocytes in the process of hyperoxia induced apoptosis and analyze the relationship between electrophysiological characteristics and morphological changes.@*METHODS@#Astrocytes were exposed to 90% hyperoxia for 12-72 h. The electrophysiological characteristics of astrocytes in each group were detected by patch clamp technique, and the morphological characteristics of astrocytes were observed at the same time. Then the same batch of astrocytes were collected, and the expression levels of caspase-1, caspase-3, gasdermin D (GSDMD) and gasdermin E (GSDME) were detected by Western blotting.@*RESULTS@#From 12 h to 72 h after hyperoxia exposure, the inward current was significantly lower than that of the control group (0.05). At each time point, the morphology of cells changed correspondingly. Western blotting showed that the expression of caspase-1 was increased significantly at 24 h and decreased significantly at 72 h after hyperoxia exposure (0.05), but began to decrease at 48 h (<0.05); GSDME increased gradually at 24 h after hyperoxia exposure (<0.05).@*CONCLUSIONS@#Under hyperoxia exposure, the ion channels of astrocytes are damaged, which can maintain the dysfunction of ion homeostasis, activate GSDME, induce the damaged cells to break away from the apoptotic pathway, and mediate the pyroptosis.


Subject(s)
Apoptosis , Astrocytes , Caspase 1 , Humans , Hyperoxia , Intracellular Signaling Peptides and Proteins , Neoplasm Proteins , Phosphate-Binding Proteins , Pyroptosis
10.
Article in Chinese | WPRIM | ID: wpr-879781

ABSTRACT

OBJECTIVE@#To investigate the role of microglial pyroptosis in hypoxic-ischemic brain damage.@*METHODS@#An oxygen-glucose deprivation/reoxygenation (OGD/R) model of rat microglial cells were cultured in vitro. Western blot was used to measure the expression of the pyroptosis-related proteins caspase-1, interleukin-1β (IL-1β), and N-terminal gasdermin D (GSDMD-N) at 0, 1, 3, 6, 12, and 24 hours after OGD/R. After the microglial cells were transfected with lentivirus-mediated silenced gasdermin D (GSDMD), immunofluorescence assay and Western blot were used to measure the transfection rate of GSDMD. Microglial cell lines were divided into three groups: normal control, negative control, and LV-sh_GSDMD (lentivirus-mediated GSDMD silencing). CCK-8 assay and LDH kit were used to observe the effect of GSDMD silencing on the viability and toxicity of microglial cells at 24 hours after OGD/R. Western blot was used to observe the effect of GSDMD silencing on the levels of caspase-1, GSDMD-N, and IL-1β in the microglial cells at 24 hours after OGD/R.@*RESULTS@#The expression levels of the pyroptosis-related proteins caspase-1, GSDMD-N, and IL-1β in microglial cells were upregulated since 0 hour after OGD/R and reached the peak levels at 24 hours. A microglial cell model of lentivirus-mediated GSDMD silencing was successfully constructed. At 24 hours after OGD/R, compared with the normal control group, the GSDMD silencing group had a significant increase in the cell viability and a significant reduction in the cytotoxicity (P<0.05), as well as significant reductions in the protein expression levels of caspase-1, GSDMD-N, and IL-1β in microglial cells (P<0.05).@*CONCLUSIONS@#Lentivirus silencing of the key substrate protein for pyroptosis GSDMD can alleviate hypoxic-ischemic brain damage, suggesting that microglial pyroptosis aggravates hypoxic-ischemic brain damage.


Subject(s)
Animals , Brain/metabolism , Intracellular Signaling Peptides and Proteins , Microglia/metabolism , Pyroptosis , Rats
11.
Arch. endocrinol. metab. (Online) ; 63(4): 385-393, July-Aug. 2019. tab, graf
Article in English | LILACS | ID: biblio-1019354

ABSTRACT

ABSTRACT Introduction Gigantism is a rare pediatric disease characterized by increased production of growth hormone (GH) before epiphyseal closure, that manifests clinically as tall stature, musculoskeletal abnormalities, and multiple comorbidities. Materials and methods Case series of 6 male patients with gigantism evaluated at the Endocrinology Service of Hospital de San José (Bogotá, Colombia) between 2010 and 2016. Results All patients had macroadenomas and their mean final height was 2.01 m. The mean age at diagnosis was 16 years, and the most common symptoms were headache (66%) and hyperhidrosis (66%). All patients had acral changes, and one had visual impairment secondary to compression of the optic chiasm. All patients underwent surgery, and 5 (83%) required additional therapy for biochemical control, including radiotherapy (n = 4, 66%), somatostatin analogues (n = 5, 83%), cabergoline (n = 3, 50%), and pegvisomant (n = 2, 33%). Three patients (50%) achieved complete biochemical control, while 2 patients showed IGF-1 normalization with pegvisomant. Two patients were genetically related and presented a mutation in the aryl hydrocarbon receptor-interacting protein (AIP) gene (pathogenic variant, c.504G>A in exon 4, p.Trp168*), fulfilling the diagnostic criteria of familial isolated pituitary adenoma. Conclusions This is the largest case series of patients with gigantism described to date in Colombia. Transsphenoidal surgery was the first-choice procedure, but additional pharmacological therapy was usually required. Mutations in the AIP gene should be considered in familial cases of GH-producing adenomas.


Subject(s)
Humans , Male , Adolescent , Young Adult , Pituitary Neoplasms/therapy , Adenoma/therapy , Gigantism/therapy , Pedigree , Pituitary Neoplasms/diagnosis , Insulin-Like Growth Factor I/analysis , Growth Hormone/blood , Adenoma/diagnosis , Retrospective Studies , Follow-Up Studies , Treatment Outcome , Sex Distribution , Colombia , Intracellular Signaling Peptides and Proteins/genetics , Growth Hormone-Secreting Pituitary Adenoma/genetics , Gigantism/diagnosis , Mutation/genetics
12.
Säo Paulo med. j ; 137(3): 255-261, May-June 2019. tab, graf
Article in English | LILACS | ID: biblio-1020958

ABSTRACT

ABSTRACT BACKGROUND: There is evidence that genetic predisposition and epigenetic alteration (e.g. DNA methylation) play major roles in lung cancer. In our genetic epidemiological studies, rs1970764 in oncogene PPP1R13L was most consistently associated with lung cancer risk. Here, we explored the role of PPP1R13L methylation in lung cancer development. DESIGN AND SETTING: Analytical cross-sectional study (45 lung cancer cases and 45 controls), conducted in China. METHODS: We investigated the DNA methylation status of 2,160 cytosine-phosphate-guanine (CpG) sites in the PPP1R13L promoter region using the EpiTYPER assay of the Sequenom MassARRAY platform. RESULTS: In the whole study group, the methylation levels of CpG-6, CpG-9, CpG-20 and CpG-21 were significantly lower and those of CpG-16 were significantly higher in cases than in controls. Among smokers, the methylation levels at five CpG sites (CpG-6, CpG-11, CpG-15, CpG-20 and CpG-21) were statistically significantly lower among cases. Among men, the methylation levels at four CpG sites (CpG-11, CpG-15, CpG-20 and CpG-21) were significantly lower among cases. Regarding smokers, the methylation levels at CpG-7.8 and CpG-21 among cases and at CpG-22 among controls were significantly lower, compared with nonsmokers. The frequency of positivity for methylation was not significantly different between lung cancer cases and controls (68.22% for cases and 71.87% for controls; P = 0.119). CONCLUSION: Our study on a Chinese population suggests that lung cancer patients have aberrant methylation status (hypomethylation tended to be more frequent) in peripheral blood leukocytes at several CpG sites in the PPP1R13L promoter region and that exposure to smoking may influence methylation status.


Subject(s)
Humans , Male , Female , Middle Aged , Aged , Repressor Proteins/genetics , DNA Methylation/genetics , Genetic Predisposition to Disease/genetics , Intracellular Signaling Peptides and Proteins/genetics , Lung Neoplasms/genetics , Case-Control Studies , Cross-Sectional Studies , Promoter Regions, Genetic
13.
Braz. j. med. biol. res ; 52(6): e8399, 2019. graf
Article in English | LILACS | ID: biblio-1011582

ABSTRACT

Imatinib is the first line of therapy for patients with metastatic or gastrointestinal stromal tumors (GIST). However, drug resistance limits the long-term effect of imatinib. Long non-coding RNAs (lncRNAs) are emerging as key players in regulating drug resistance in cancer. In this study, we investigated the association between lncRNA CCDC26 and IGF-1R in GIST and their involvement in drug resistance. Considering the key role of lncRNAs in drug resistance in cancer, we hypothesized that IGF-1R is regulated by lncRNAs. The expression of a series of reported drug resistance-related lncRNAs, including CCDC26, ARF, H19, NBR2, NEAT1, and HOTAIR, in GIST cells treated with imatinib H19 was examined at various time-points by qRT-PCR. Based on our results and published literature, CCDC26, a strongly down-regulated lncRNA following imatinib treatment, was chosen as our research target. GIST cells with high expression of CCDC26 were sensitive to imatinib treatment while knockdown of CCDC26 significantly increased the resistance to imatinib. Furthermore, we found that CCDC26 interacted with c-KIT by RNA pull down, and that CCDC26 knockdown up-regulated the expression of IGF-1R. Moreover, IGF-1R inhibition reversed CCDC26 knockdown-mediated imatinib resistance in GIST. These results indicated that treatments targeting CCDC26-IGF-1R axis would be useful in increasing sensitivity to imatinib in GIST.


Subject(s)
Humans , Receptors, Somatomedin/genetics , Drug Resistance, Neoplasm , Intracellular Signaling Peptides and Proteins/genetics , RNA, Long Noncoding/genetics , Imatinib Mesylate/pharmacology , Antineoplastic Agents/pharmacology , Signal Transduction , Down-Regulation/genetics , Gene Expression Regulation, Neoplastic , Receptors, Somatomedin/metabolism , Receptor, IGF Type 1 , Apoptosis , Cell Line, Tumor , Intracellular Signaling Peptides and Proteins/metabolism , RNA, Long Noncoding/metabolism , Flow Cytometry
14.
Experimental Neurobiology ; : 628-641, 2019.
Article in English | WPRIM | ID: wpr-763784

ABSTRACT

Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5) has been reported to play critical roles in the proliferation of various cancer cells. However, the roles of LGR5 in brain tumors and the specific intracellular signaling proteins directly associated with it remain unknown. Expression of LGR5 was first measured in normal brain tissue, meningioma, and pituitary adenoma of humans. To identify the downstream signaling pathways of LGR5, siRNA-mediated knockdown of LGR5 was performed in SH-SY5Y neuroblastoma cells followed by proteomics analysis with 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE). In addition, the expression of LGR5-associated proteins was evaluated in LGR5-inhibited neuroblastoma cells and in human normal brain, meningioma, and pituitary adenoma tissue. Proteomics analysis showed 12 protein spots were significantly different in expression level (more than two-fold change) and subsequently identified by peptide mass fingerprinting. A protein association network was constructed from the 12 identified proteins altered by LGR5 knockdown. Direct and indirect interactions were identified among the 12 proteins. HSP 90-beta was one of the proteins whose expression was altered by LGR5 knockdown. Likewise, we observed decreased expression of proteins in the hnRNP subfamily following LGR5 knockdown. In addition, we have for the first time identified significantly higher hnRNP family expression in meningioma and pituitary adenoma compared to normal brain tissue. Taken together, LGR5 and its downstream signaling play critical roles in neuroblastoma and brain tumors such as meningioma and pituitary adenoma.


Subject(s)
Brain , Brain Neoplasms , Cell Proliferation , Dermatoglyphics , Electrophoresis, Polyacrylamide Gel , GTP-Binding Proteins , Heterogeneous-Nuclear Ribonucleoproteins , Humans , Intracellular Signaling Peptides and Proteins , Meningioma , Neuroblastoma , Pituitary Neoplasms , Proteomics
15.
Article in Chinese | WPRIM | ID: wpr-774567

ABSTRACT

The aim of this study was to observe the effect of baicalin on the growth state of attention deficit hyperactivity disorder animal model and its regulation on Ca MKⅡand ERK1/2.In the present study,a total of 40 SHR rats were randomly divided into model group,methylphenidate hydrochloride group,and low,medium,and high dose baicalin groups,with 8 rats in each group.Eight WKYrats were selected as a normal control group.The methylphenidate hydrochloride group(0.07 g·L~(-1))and the low(3.33 g·L~(-1)),medium(6.67 g·L~(-1)),and high dose(10 g·L~(-1))baicalin groups received corresponding drugs by gavage administration according to the body weight(0.015 m L·g~(-1)),while the normal group and the model group received the same volume of normal saline by gavage.Thegavage administration lasted for 4 weeks,twice a day.The body weight of the rats and the amount of remaining feed were weighed daily,and the growth state of the rats was statistically evaluated weekly.Percoll density gradient centrifugation was used to prepare brain synaptosomes and an electron microscope was used to observe their structures.The Ca MKⅡand ERK1/2 protein and mRNA expression levels were detected with Western blot and Real-time PCR methods,respectively.RESULTS: showed that baicalin did not affect the normal eating and weight gain of rats,and the weight gain of rats was even more significant than that in the normal group(P<0.05).In the study of its effects on Ca MKⅡand ERK1/2 protein expression in rat synaptosomes,the expression of both proteins in each drug-administered group was higher than that in the model group(P<0.05);besides,the expression levels of Ca MKⅡand ERK1/2 protein were significantly increased in both baicalin high dose group and the methylphenidate hydrochloride group(P<0.05).The relative expression of Ca MKⅡand ERK1/2 mRNA in synaptosome was detected by PCR.The results showed that medium and high doses of baicalin and methylphenidate hydrochloride significantly increased the relative expression of Ca MKⅡand ERK1/2 mRNA in synaptosomes of SHR rats(P<0.05).In conclusion,baicalin does not affect the normal growth and development of SHR rats,so it is safe for administration.Both baicalin and methylphenidate hydrochloride could up-regulate the relative expression of Ca MKⅡand ERK1/2 in mRNA and protein,and the pharmacodynamic stability of baicalin is in a dose-dependent manner to certain extent.


Subject(s)
Animals , Attention Deficit Disorder with Hyperactivity , Disease Models, Animal , Flavonoids , Intracellular Signaling Peptides and Proteins , MAP Kinase Signaling System , Protein-Serine-Threonine Kinases , Rats , Rats, Inbred SHR , Rats, Inbred WKY
16.
Article in Chinese | WPRIM | ID: wpr-781314

ABSTRACT

OBJECTIVE@#To carry out prenatal diagnosis for a women with Branchio-oto-renal syndrome by using chromosomal microarray analysis (CMA).@*METHODS@#Peripheral blood chromosomal karyotyping and CMA were used to analyze the gravida with an abnormal phenotype. Pathological copy number variants (CNVs) were validated in other members of the family members and her fetus.@*RESULTS@#The gravida and her daughter both had Branchio-oto-renal syndrome and a 8q13.3 microdeletion encompassing the EYA1 gene. The same microdeletion was also found in the fetus. No phenotypic or genotypic anomaly was found with other members of the family.@*CONCLUSION@#Mutation of the EYA1 gene probably underlies the Branchio-oto-renal syndrome in this family, which is consistent with an autosomal dominant inheritance.


Subject(s)
Branchio-Oto-Renal Syndrome , Diagnosis , Genetics , Female , Humans , Intracellular Signaling Peptides and Proteins , Genetics , Nuclear Proteins , Genetics , Pedigree , Pregnancy , Prenatal Diagnosis , Protein Tyrosine Phosphatases , Genetics
17.
Article in English | WPRIM | ID: wpr-785903

ABSTRACT

Macrophages play essential roles in innate immune responses by producing various immune mediators. Therefore, modulating macrophage function is an attractive strategy to treat immune disorders. Aralia cordata var. continentalis (AC), known as “Dokwhal” in Korea, possesses various biological and medicinal functions, including immunomodulation. The present study investigated the effect of the hot water extract of AC (HAC) on RAW264.7 murine macrophages. When these cells were treated with HAC, nitric oxide production and inducible nitric oxide synthase expression was induced dose-dependently. In addition, HAC treatment triggered the secretion of innate immune cytokines, such as TNF-α and IL-6. Phagocytosis, measured by FITC-dextran internalization showed that HAC stimulated the phagocytic activity of macrophages. Furthermore, HAC promoted the production of reactive oxygen species in RAW264.7 cells, determined by CM-H2DCFDA. In addition, the immunoblot analysis of intracellular signaling proteins revealed that NF-kB and MAPK signaling pathways, which are important signaling mediators of inflammation, are upregulated by HAC. In conclusion, these findings suggested that HAC can stimulate macrophage activity, and NF-kB and MAPK signaling pathways might be involved in the immunostimulatory effects of HAC.


Subject(s)
Aralia , Cytokines , Immune System Diseases , Immunity, Innate , Immunomodulation , Inflammation Mediators , Interleukin-6 , Intracellular Signaling Peptides and Proteins , Korea , Macrophages , NF-kappa B , Nitric Oxide , Nitric Oxide Synthase Type II , Phagocytosis , Reactive Oxygen Species , Water
18.
Article in Chinese | WPRIM | ID: wpr-771962

ABSTRACT

OBJECTIVE@#To identify potential mutation in a child clinically diagnosed as Noonan syndrome and to provide genetic counseling and prenatal diagnosis for his family.@*METHODS@#Genomic DNA was extracted from peripheral blood samples of the patient and his parents, and amniotic fluid was taken from the mother during the second trimester. Next generation sequencing (NGS) was used to screen potential mutations from genomic DNA. Suspected mutation was verified by Sanger sequencing.@*RESULTS@#A heterozygous c.4A>G (p.Ser2Gly) mutation of the SHOC2 gene was identified in the patient but not among other family members including the fetus.@*CONCLUSION@#The Noonan syndrome is probably caused by the c.4A>G mutation of the SHOC2 gene. NGS is helpful for the diagnosis of complicated genetic diseases. SHOC2 gene mutation screening is recommended for patient suspected for Noonan syndrome.


Subject(s)
Child , Female , Genetic Testing , High-Throughput Nucleotide Sequencing , Humans , Intracellular Signaling Peptides and Proteins , Mutation , Noonan Syndrome , Pregnancy , Prenatal Diagnosis
19.
Article in Chinese | WPRIM | ID: wpr-773545

ABSTRACT

OBJECTIVE@#To study the value of plasma miRNA23-a and miRNA-451 as potential biomarkers for early diagnosis of non-small cell lung cancer (NSCLC).@*METHODS@#Fifty patients with NSCLC and 50 healthy control subjects were recruited for testing the plasma levels of miRNA23-a and miRNA-451 and their expression levels in the tumor tissues using qRT-PCR. The correlations of the plasma levels of miRNA23-a and miRNA-451 with their expressions in the tumor tissues were analyzed. The diagnostic power of CEA, miRNA23-a and miRNA-451 for NSCLC was evaluated using the receiver-operating characteristics (ROC) curves and the area under the ROC curves (AUC). In the NSCLC cell line A549, we tested the effect of inhibition of miRNA-23a and miRNA-451 on the expression levels of SPRY2 and MIF mRNA using qRT-PCR.@*RESULTS@#The expression levels of miRNA-23a and miRNA-451 in NSCLC tissues was significantly associated with smoking, tumor size, lymph node metastasis and TNM stage ( < 0.05). Compared with those in the control group, miRNA-23a level was significantly increased while miRNA-451 was significantly down-regulated in the tumor tissues and plasma of NSCLC patients. The plasma levels of miRNA-23a and miRNA-45 were strongly correlated with their expression levels in the tumor tissues. ROC analysis showed that for the diagnosis of NSCLC, the AUC, sensitivity and specificity of either miRNA-23a or miRNA-451 were significantly higher than those of CEA ( < 0.05). The combination of miRNA23-a and miRNA-451 markedly improved the AUC (0.900), sensitivity (78%) and specificity (86%) for the diagnosis. In A549 cells, inhibition of miRNA23-a and miRNA-451 resulted in significantly increased expression levels of SPRY2 mRNA and MIF mRNA, respectively.@*CONCLUSIONS@#miRNA-23a and miRNA-451 can be used as potential biomarkers for early diagnosis of NSCLC, and their combined detection can be more effective for the diagnosis.


Subject(s)
Biomarkers, Tumor , Carcinoma, Non-Small-Cell Lung , Genetics , Case-Control Studies , Early Detection of Cancer , Humans , Intracellular Signaling Peptides and Proteins , Lung Neoplasms , Genetics , Membrane Proteins , MicroRNAs , ROC Curve
20.
Article in Chinese | WPRIM | ID: wpr-813289

ABSTRACT

To explore the clinical significance of the altered expression of polycomb group (PcG)-associated protein RYBP in hepatocellular carcinoma (HCC) specimens.
 Methods: The expression levels of RYBP in tumor tissues and adjacent normal tissues in 77 HCC cases were detected by immunohistochemistry (IHC), and the relationships between RYBP expression levels and HCC clinicopathological characteristics, five-year survival rates or prognosis of HCC patients were analyzed.
 Results: RYBP expression level was significantly decreased in HCC tumor tissues than that in the adjacent normal tissues (P<0.05). The expression levels of RYBP in HCC specimens were highly correlated with HBsAg, ALT, GGT, Type III procollagen, tumor size, distant metastasis, and tumor differentiation (P<0.05). The RFS and OS for patients with RYBP-low expression were markedly lower than those with RYPB-high expression (P<0.05). Both age and RYBP expression level were protective factors for RFS, while GGT, lymph node metastasis, TNM stage, tumor differentiation and tumor size were risk factors for RFS (P<0.05). As to OS, RYBP expression level was a protective factor, while tumor number, ALT, GGT, AFP, pCEA, lymph node metastasis, TNM stage, tumor differentiation and tumor size were risk factors (P<0.05). The age, GGT, lymph node metastasis and TNM stage were independent prognostic factors for RFS (P<0.05), and both lymph node metastasis and TNM stage were independent risk factors for OS (P<0.05). Comparing to serum alpha fetoprotein (AFP) level, RYBP expression level in tumor tissues was applied to predict the prognosis of HCC patients more accurately.
 Conclusion: PcG associated protein RYBP displays a reduced expression in HCC tissues, which is related to poor prognosis of HCC patients. It might be a promising therapeutic target for HCC treatment.


Subject(s)
Biomarkers, Tumor , Carcinoma, Hepatocellular , Humans , Immunohistochemistry , Intracellular Signaling Peptides and Proteins , Metabolism , Liver Neoplasms , Lymphatic Metastasis , Prognosis
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