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Background: The canonical Wnt signal transduction or the Wnt/ß-catenin pathway plays a crucial role in both carcinogenesis and development of animals. Activation of the Gαq class of Gα proteins positively regulates Wnt/ß-catenin pathway, and expression of Gαq in human embryonic kidney 293 (HEK293T) cells or Xenopus oocytes leads to the inhibition of glycogen synthase kinase-3 beta and cellular accumulation of ß-catenin. This study investigated whether Gαq-mediated cellular accumulation of ß-catenin could affect the transcriptional activity of this protein. Methods: HEK-293T and HT-29 cells were used for cell culture and transfection. Protein localization and quantification were assessed by using immunofluorescence microscopy, cell fractionation assay, and Western blotting analysis. Gene expression at the transcription level was examined by quantitative reverse transcriptase/real-time PCR method. Results: Transcription of two cellular ß-catenin target genes (c-MYC and CCND1) and the ß-catenin/ T-cell factor reporter luciferase gene (TopFlash plasmid) significantly increased by Gαq activation. The Gαq-mediated increase in the expression level of the ß-catenin-target genes was sensitive to the expression of a minigene encoding a specific Gαq blocking peptide. The results of cell fractionation and Western blotting experiments showed that activation of Gαq signaling increased the intracellular ß-catenin protein level, but it blocked its membrane localization. Conclusion: Our results reveal that the Gαq-dependent cellular accumulation of ß-catenin can enhance ß-catenin transcriptional activity.
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Via de Sinalização Wnt , beta Catenina , Animais , Humanos , beta Catenina/genética , Células Epiteliais , Células HEK293 , Transcrição Gênica , Via de Sinalização Wnt/genética , XenopusRESUMO
Background: Fibrillin is one of the extracellular matrix glycoproteins and participates in forming microfibrils found in many connective tissues. The microfibrils enable the elasticity and stretching properties of the ligaments and support connective tissues. There are three isoforms of fibrillin molecules identified in mammals: fibrillin 1 (FBN1), fibrillin 2 (FBN2), and fibrillin 3. Objective: Multiple studies have shown that mutations in these genes or changes in their expression levels can be related to various diseases, including cancers. In this study, we focus on reviewing the role of the fibrillin family in multiple cancers. Methods and Results: We performed a comprehensive literature review to search PubMed and Google Scholar for studies published so far on fibrillin gene expression and its role in cancers. In this review, we have focused on the expression of FBN1 and FBN2 genes in cancers such as the lung, intestine, ovary, pancreatic ductal, esophagus, and thyroid. Conclusion: Altogether various studies showed higher expression of fibrillins in different tumor tissues correlated with the patient's survival. However, there are controversial findings, as some other cancers showed hypermethylated FBN promoters with lower gene expression levels.
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BACKGROUND: Adenosine, a purine nucleoside, plays an important function in the pathogenesis of cancer through interaction with the cell surface G protein-coupled adenosine receptors. It is important to determine the expression pattern of these receptors in different cancers. Previously in our lab, we found up-regulation of A1 adenosine receptor (AR) in lung tumors playing as a putative target for cancer cell inhibition, and here we aimed to investigate the significance of other adenosine receptor isoforms (A2aAR, A2bAR, and A3AR). METHODS: In this study, first of all, we evaluated the adenosine receptors gene expression in the bioinformatics database (GENT2). Then the genes expression was measured experimentally in the 20 lung cancer tumor tissues in comparison to the matched tumor-adjacent normal tissue (as control). The mRNA expression of receptors was evaluated by real-time PCR. The tumors were categorized by the tumor size and the gene expression change was evaluated. RESULTS: The experimental results indicated a significant increase in A2aAR (p value = .021) and A3AR (p value = .01) expression in lung tumor tissues compared to the adjacent tumor margins which were in accordant to bioinformatics analysis. We found a non-significant increase in A2bAR expression; however, when comparing the patients according to the tumor size, our data showed that the expression of A2bAR adenosine receptor in patients with smaller lung tumor sizes was higher than the other group (p = .011). CONCLUSION: The results of this study showed that adenosine receptors A3AR, and A2aAR are highly expressed in lung tumors relative to tumor-adjacent normal tissue. We suggest that overexpression of adenosine receptors in lung cancer is due to their regulatory role in various aspects of lung cancer.
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Neoplasias Pulmonares , Receptores Purinérgicos P1 , Humanos , Receptores Purinérgicos P1/genética , Adenosina/farmacologia , Adenosina/fisiologia , Neoplasias Pulmonares/genética , Expressão GênicaRESUMO
BACKGROUND: Esophageal cancer (EC) is the sixth leading cause of cancer-related death, despite many advances in treatment, the survival of patients still remains poor. In recent years, the N6-methyladenosine (m6A) has been introduced as one of the most important modifications at the epitranscriptome level, with an important role in the mRNA regulation in various diseases, such as cancers. The m6A is regulated by different factors, including FTO as a demethylase. The m6A modification, especially through FTO overexpression has an oncogenic role in different cancer types such as EC. Recent studies showed that entacapone, a catechol-o-methyl transferase (COMT) inhibitor currently applied for Parkinson's disease, can inhibit FTO enzyme. AIMS: In this study, we aimed to investigate the effect of entacapone as an FTO inhibitor on the m6A level and also apoptosis and cell cycle response in KYSE-30 and YM-1 of esophageal squamous cancer cell (ESCC) lines. METHODS: Cell toxicity and IC50 of entacapone were evaluated using The MTT assay in YM-1 and KYSE-30 cells. Cells were treated into two groups: DMSO (control) and entacapone (mean IC50 ). Total RNA was extracted, and m6A levels were measured via the ELISA method. Subsequently, the apoptosis and cell cycle dys-regulation were detected by annexin-V-FITC/PI staining and PI staining via flow cytometry. RESULTS: Entacapone has the cytotoxicity effect on both esophageal cancer cell lines compared to normal PBMC cells. As well, entacapone treatment (140 µM) can induce apoptosis (KYSE-30: 50%. YM-1:22.6%) and has a modulatory effect on cell cycle progression in both YM-1 and KYSE-30 cells (p-value<.05). However, no significant difference in the m6A concentration was observed. CONCLUSION: Our findings suggested that entacapone has the inhibitory effect on ESCC cell lines through induction of the apoptosis and modulation of the cell cycle without toxicity on the normal PBMC.
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Catecol O-Metiltransferase , Neoplasias Esofágicas , Humanos , Catecol O-Metiltransferase/farmacologia , Leucócitos Mononucleares/metabolismo , Neoplasias Esofágicas/tratamento farmacológico , Apoptose , Ciclo Celular , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/metabolismoRESUMO
BACKGROUND: SOX2OT is a novel cancer associated long non-coding RNA (LncRNA) with higher expression in variable tumor tissues, including esophageal squamous cell carcinoma (ESCC). It also plays an important function in embryonic neuronal development. Regarding its function in both stemness and carcinogenesis, here, we aimed to investigate its expression and function in tumorspheres of the esophagus using the RNAi method. MATERIAL & METHODS: Two esophageal squamous cancer cells (ESCC): KYSE30 and YM1 cells were used for sphere enrichment. Cells were transfected with SOX2OT targeting and control siRNA. The size and the number of spheres were measured using light microscopy. Gene expression of the pluripotency genes was measured by qRT-PCR and docetaxel chemoresistance was assessed by MTS viability assay. RESULTS: Our findings showed that ESCC tumorspheres overexpress SOX2OT gene along with other stemness genes (SOX2, OCT4A, and Nanog) compared to their original cancer cells. RNAi experiments indicated that SOX2OT knockdown can suppress the stemness-related gene expression, sphere formation ability (both size and number), and docetaxel resistance as three of the main cancer stem cell characteristics of tumorspheres. CONCLUSION: Altogether our results showed the regulatory role of SOX2OT in pluripotency and stemness in ESCC tumorspheres. Our results suggest a potential application of SOX2OT inhibition in combination with docetaxel for ESCC inhibition in vitro.
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Purpose: Esophageal squamous cell carcinoma (ESCC) is a highly aggressive cancer. The main cause of death in ESCC is related to relapse, metastasis, and resistance to cancer therapy. Recent studies have shown that a minor subset of cancer cells, known as cancer stem cells (CSCs), are responsible for tumor formation initiation and cancer progression. Understanding the genes associated with CSCs and metastasis can help in targeted cancer therapy. The aim of this study was to assess the expression of LAMB3 and TOP2A metastasis-associated genes in CSCs and adherent cells in the xenograft mouse model. Methods: Esophageal CSCs were enriched by the sphere formation method. The expression level of LAMB3 and TOP2A genes were evaluated in spheres and adherent cells in vitro by qRT-PCR. A xenograft mouse model was established to investigate the tumorigenesis and metastasis potential by subcutaneous and tail vein injection of CSCs and adherent YM-1 cells. Consequently, LAMB3 and TOP2A expression at the mRNA level was assessed in tumors. Immunohistochemistry was also used to evaluate the LAMB3 expression at the protein level in tumors. Results: CSCs-derived tumor was developed more quickly than the adherent cells-derived tumor. LAMB3 at mRNA and protein level was significantly down-regulated in sphere-derived tumor compared with adherent cells-derived tumor (P value <0.05). TOP2A expression was almost similar in both sphere cells and adherent cells and there was no significant difference. Conclusion: we concluded that YM-1 spheres have CSCs characteristics in vitro with high capability of tumorigenicity in vivo. Our results were also shown that the LAMB3 expression was decreased in YM-1 spheres suggesting LAMB3 association with sphere formation.
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Systemic sclerosis (SSc) is a chronic autoimmune disease, featuring fibrosis in multiple organs. The serum from SSc patients contain inflammatory mediators, contributing to SSc pathogenesis and could be used to develop cell culture models. Here, we compared the fibrotic effects of serum samples from SSc patients with TGFß1 on human dermal fibroblasts (HDFs). HDF cells were cultured in four different culture media supplementations; 10% SSc serum, 10% healthy human serum, 10% fetal bovine serum or 10% FBS supplemented with 10 ng/Ml human TGFß. The collagen content in cell layers was measured by spectrophotometric Picro-Sirius red staining. The mRNA expression of α-SMA, COL I and III, TGFß1, arginase and E-Cadherin genes were determined by real time RT-PCR. TGF-ß1 levels in cell culture supernatants were measured using ELISA. Cell layer collagen content was significantly increased following TGF-ß1 treatment, compared with FBS group and SSc serum treatment in comparison with healthy controls. Although not statistically significant, the mRNA expression of α-SMA, COLI and III, TGFß1, and arginase increased upon TGF-ß1 treatment in comparison with FBS group, and in SSc serum treatment group in comparison with healthy controls. E-Cadherin decreased following TGF-ß1 treatment and SSc serum treatment in comparison with their counterparts. TGF-ß1 levels increased in cell culture supernatants of HDF cells exposed to TGF-ß1 and SSc serum. An in vitro model of SSc serum-induced fibrosis using human HDF cells was evaluated in comparison to the TGF-ß1 fibrosis induced model and data suggested that it may be used in documenting the role of pro-fibrotic factors in serum or plasma from SSc patients.
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OBJECTIVES: Fibrosis is a chronic inflammation caused by the loss of innate compensational mechanisms. Naringin (NR) is a flavonoid with antineoplastic and anti-inflammatory effects. Here, we aimed to investigate the antifibrotic effects of NR and underlying mechanisms in a Hypochlorous acid (HOCl)-induced mouse model of skin fibrosis. MATERIALS AND METHODS: A total of 24 six-week-old female BALB/c mice were randomly allocated into five groups: HOCl, Sham, PBS, HOCl + NR and DMSO and selected skin regions were treated for 6 weeks, until sacrifice. The histopathologic and collagenesis of skin resections were analyzed using H&E and PR staining. The mRNA levels of COL1, COL3 and αSMA genes were quantified. Serum samples were also used to evaluate TGF-ß levels and LDH activity. RESULTS: HOCl could increase the relative collagen content, while NR administration on HOCl-treated biopsies decreased collagenesis. COL1, COL3 and αSMA mRNA levels were significantly increased among HOCl-treated skin samples, while NR treatment could decrease these mRNA levels of genes to the extent equal to the levels in the Sham group. Similarly, Naringin-treated samples could decrease TGF-ß levels. CONCLUSIONS: We demonstrated that Naringin could exert protective effects against fibrotic complications of HOCL in skin tissue in vivo, by reducing the collagenesis and decreasing the levels of fibrosis-associated genes.
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Flavanonas , Dermatopatias , Animais , Feminino , Camundongos , Anti-Inflamatórios/farmacologia , Colágeno/farmacologia , Dimetil Sulfóxido , Modelos Animais de Doenças , Fibrose , Flavanonas/farmacologia , Ácido Hipocloroso/efeitos adversos , Camundongos Endogâmicos BALB C , RNA Mensageiro , Fator de Crescimento Transformador beta , Dermatopatias/induzido quimicamente , Dermatopatias/tratamento farmacológicoRESUMO
BACKGROUND: Genetic variations of aryl hydrocarbon receptor (AHR) pathway genes could influence the imbalanced immune response to xenobiotics. Therefore, we aimed to investigate the polymorphism of AHR pathway genes in systemic lupus erythematosus (SLE) patients in association with smoking. METHODS: Genomic DNA from patients (N = 107) and controls (N = 105) of a population from northeast of Iran was used for genotyping of CYP1A1 T>C (rs4646903) and AHRR C>G (rs2292596) variants. The SLEDAI score and smoking status of the patients were registered. The AHR activity was estimated by CYP1A1 and CYP1B1 gene expression in peripheral blood mononuclear cells (PBMC). RESULTS: The C allele in rs4646903 (odds ratio [OR] = 2.67) and G allele in rs2292596 (OR = 1.79) SNPs were significantly associated with the increased risk of SLE. The AHR pathway was more active in high-risk CYP1A1/AHRR: C/G haplotype. The most severe disease was observed in smoker patients with high-risk haplotype and both smoking (Exp (ß) = 9.5) and high-risk CYP1A1/AHRR (C/G) haplotype (Exp (ß) = 3.7) can significantly increase the likelihood of having severe (SLEDAI ≥ 20) SLE disease activity. CONCLUSION: Our findings indicated the association of xenobiotic-metabolizing genes (CYP1A1, AHRR) polymorphisms with the susceptibility to SLE and disease severity regarding the smoking background, suggesting the interaction of gene and environmental risk factors in SLE pathogenesis.
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Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Fumar Cigarros , Citocromo P-450 CYP1A1/genética , Predisposição Genética para Doença , Lúpus Eritematoso Sistêmico/genética , Proteínas Repressoras/genética , Adulto , Alelos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Estudos de Casos e Controles , Feminino , Interação Gene-Ambiente , Variação Genética , Genótipo , Haplótipos , Humanos , Lúpus Eritematoso Sistêmico/patologia , Masculino , Proteínas Repressoras/metabolismo , Índice de Gravidade de Doença , Xenobióticos/metabolismoRESUMO
Molecular knowledge regarding the primary esophageal achalasia is essential for the early diagnosis and treatment of this neurodegenerative motility disorder. Therefore, there is a need to find the main microRNAs (miRNAs) contributing to the mechanisms of achalasia. This study was conducted to determine some patterns of deregulated miRNAs in achalasia. This case-control study was performed on 52 patients with achalasia and 50 nonachalasia controls. The miRNA expression profiling was conducted on the esophageal tissue samples using the next-generation sequencing (NGS). Differential expression of miRNAs was analyzed by the edgeR software. The selected dysregulated miRNAs were additionally confirmed using the quantitative reverse transcription polymerase chain reaction (qRT-PCR). Fifteen miRNAs were identified that were significantly altered in the tissues of the patients with achalasia. Among them, three miRNAs including miR-133a-5p, miR-143-3p, and miR-6507-5p were upregulated. Also, six miRNAs including miR-215-5p, miR-216a-5p, miR-216b-5p, miR-217, miR-7641 and miR-194-5p were downregulated significantly. The predicted targets for the dysregulated miRNAs showed significant disease-associated pathways like neuronal cell apoptosis, neuromuscular balance, nerve growth factor signaling, and immune response regulation. Further analysis using qRT-PCR showed significant down-regulation of hsa-miR-217 (p-value = 0.004) in achalasia tissue. Our results may serve as a basis for more future functional studies to investigate the role of candidate miRNAs in the etiology of achalasia and their application in the diagnosis and probably treatment of the disease.
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The pandemic of coronavirus disease 2019 (COVID-19), with rising numbers of patients worldwide, presents an urgent need for effective treatments. To date, there are no therapies or vaccines that are proven to be effective against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Several potential candidates or repurposed drugs are under investigation, including drugs that inhibit SARS-CoV-2 replication and block infection. The most promising therapy to date is remdesivir, which is US Food and Drug Administration (FDA) approved for emergency use in adults and children hospitalized with severe suspected or laboratory-confirmed COVID-19. Herein we summarize the general features of SARS-CoV-2's molecular and immune pathogenesis and discuss available pharmacological strategies, based on our present understanding of SARS-CoV and Middle East respiratory syndrome coronavirus (MERS-CoV) infections. Finally, we outline clinical trials currently in progress to investigate the efficacy of potential therapies for COVID-19.
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Imunidade Adaptativa , Betacoronavirus/fisiologia , Infecções por Coronavirus/patologia , Pneumonia Viral/patologia , Anti-Inflamatórios/uso terapêutico , Antivirais/uso terapêutico , Betacoronavirus/isolamento & purificação , COVID-19 , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/terapia , Infecções por Coronavirus/virologia , Humanos , Imunoterapia , Coronavírus da Síndrome Respiratória do Oriente Médio/isolamento & purificação , Coronavírus da Síndrome Respiratória do Oriente Médio/fisiologia , Fases de Leitura Aberta/genética , Pandemias , Pneumonia Viral/imunologia , Pneumonia Viral/terapia , Pneumonia Viral/virologia , SARS-CoV-2RESUMO
INTRODUCTION: The aim of this study was to assess the effects of Rubus anatolicus on glucose metabolism in HepG2, CRI-D2 and C2C12 cell lines. MATERIALS AND METHODS: R. anatolicus was collected in Golestan province, Iran. Three different cell lines HepG2 (human liver cell), CRI-D2 (mice pancreatic cell) and C2C12 (rat myoblast) were used for cell culture experiments. Cell viability was measured using MTT assay. Cells were treated with various concentrations of the extract (6.25-400 µg/mL) and then the extracellular glucose level and intracellular glycogen content were measured using colorimetric methods. The insulin level of the culture medium was measured using the ELISA method. RESULTS: Our findings showed that R. anatolicus extract enhances glucose uptake and consumption by all three cell lines. The R. anatolicus extract exposure also elevated cellular glycogen content in HepG2 and C2C12 cells (for 200 and 100 µg/mL) significantly. We found a significant increase in glucose uptake and consequently higher stimulation of insulin secretion in CRI-D2 cell pancreatic cells treated with R. anatolicus extract. CONCLUSION: The R. anatolicus appears to activate glucose uptake and cellular glycogen synthesis probably by activating the glycogenesis or inhibition of glycogenolysis pathways. The extract enhances insulin secretion in the pancreatic cells by increased glucose uptake.
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The field of gene therapy is striving more than ever to define a path to the clinic and the market. Twenty gene therapy products have already been approved and over two thousand human gene therapy clinical trials have been reported worldwide. These advances raise great hope to treat devastating rare and inherited diseases as well as incurable illnesses. Understanding of the precise pathomechanisms of diseases as well as the development of efficient and specific gene targeting and delivery tools are revolutionizing the global market. Currently, human cancers and monogenic disorders are indications number one. The elevated prevalence of genetic disorders and cancers, clear gene manipulation guidelines and increasing financial support for gene therapy in clinical trials are major trends. Gene therapy is presently starting to become commercially profitable as a number of gene and cell-based gene therapy products have entered the market and the clinic. This article reviews the history and development of twenty approved human gene and cell-based gene therapy products that have been approved up-to-now in clinic and markets of mainly North America, Europe and Asia.
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OBJECTIVE: Periodontitis is an inflammatory disease that is a result of the interaction between pathogenic bacteria and host immune response. Genetic alterations in interleukin-10 (IL-10) gene may be associated with the increased risk of periodontitis. We investigated the association between genotype and haplotype frequencies of IL-10 gene polymorphisms and susceptibility to periodontitis in an Iranian population. METHODS: In this case-control study, a total of 64 patients with periodontitis and 128 healthy subjects were recruited. The PCR-RFLP technique was used to detect IL-10 promoter genotypes at the positions of -1082 (G/A), -819 (C/T), and -592 (C/A) in association with the susceptibility to severe chronic periodontitis. RESULTS: Regarding IL-10 -592 (C/A) and IL-10 -819 (C/T) alleles and genotypes, no significant association was observed between the risk of periodontitis and genotype frequencies. However, the frequency of GG genotype in IL-10 -1082 (G/A) polymorphic region was higher in normal subjects and was associated with the decreased risk of periodontitis under recessive model [OR = 2.89, 95% CI (0.99-8.43), p = 0.039]. Haplotype analysis revealed a significantly higher presence of H7 (AGC; -592/-1082/-819) [OR = 97.74, 95% CI (95.52-99.96), p < 0.0001]. CONCLUSION: IL-10 -1082(G/A) polymorphism and AGC (-592/-1082/-819) haplotype could be associated with the susceptibility to chronic periodontitis.
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Periodontite Crônica/genética , Predisposição Genética para Doença/genética , Interleucina-10/genética , Alelos , Estudos de Casos e Controles , Periodontite Crônica/etnologia , Frequência do Gene , Predisposição Genética para Doença/etnologia , Genótipo , Humanos , Irã (Geográfico)/epidemiologia , Polimorfismo Genético , Polimorfismo de Nucleotídeo ÚnicoRESUMO
Systemic lupus erythematosus (SLE) is a chronic autoimmune disease with unknown aetiology. According to the role of interleukin 10 (IL10) in SLE pathogenesis, the genetic alterations in its promoter region could be associated with elevated IL10 levels and exacerbated disease. Here, we investigated the association of genotype and haplotype frequencies of three IL10 gene promoter polymorphisms with susceptibility to SLE, IL10 plasma levels and disease activity of patients in an Iranian population. A total of 116 SLE patients and 131 healthy subjects were enrolled. The PCR-RFLP technique was used to detect IL10 promoter genotypes at the positions of -1082 (G/A), -819 (C/T) and -592 (C/A) in association with IL10 plasma levels and SLEDAI scores. The GG genotype of -1082 polymorphism was associated with the increased risk of SLE [OR = 2.65, 95% CI (1.21-5.82), p-value = 0.046]. The CC genotype in -819 region was associated with SLE susceptibility [OR = 3.38, 95% CI (1.26-9.07), p-value = 0.034] and C allele was introduced as risk allele [OR = 1.86, 95% CI (1.15-3.01), p-value = 0.009] in this region. IL10 plasma levels were overexpressed in CC genotype carriers of -592 SNP and decreased in AA genotype carriers of -1082. IL10 was also increased in SLE patients with CGT (-592/-1082/-819) haplotype. The SLEDAI score was higher among CC genotype carriers at the position of -592 and TT genotype carriers at the region of -819. SLEDAI was also elevated among patients with CGC (-592/-1082/-819) and CAC (p = 0.011) haplotypes. The present study suggests that the IL10 -819(C/T), -1082(G/A) and -592(C/A) polymorphisms and the haplotypes are associated with SLE susceptibility, increased disease activity and elevated IL10 levels. While this is the first time to report such an association in an Iranian population, further studies are needed to confirm these findings.
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Estudos de Associação Genética , Predisposição Genética para Doença , Interleucina-10/genética , Lúpus Eritematoso Sistêmico/genética , Adulto , Alelos , Feminino , Genótipo , Haplótipos , Voluntários Saudáveis , Humanos , Interleucina-10/sangue , Irã (Geográfico)/epidemiologia , Lúpus Eritematoso Sistêmico/sangue , Lúpus Eritematoso Sistêmico/epidemiologia , Lúpus Eritematoso Sistêmico/fisiopatologia , Masculino , Polimorfismo de Nucleotídeo Único/genética , Regiões Promotoras Genéticas/genéticaRESUMO
BACKGROUND: SOX2 overlapping transcript (SOX2OT) is a long non-coding RNA, over-expressed in human tumor tissues and embryonic cells. Evidences support its function in the cell cycle; however there is no clear mechanism explaining its function in cell proliferation regulation. Here we investigated cancer cell response to SOX2OT knockdown by RNA sequencing. METHODS: SOX2OT expression was inhibited by siRNA in two cancer cell lines (A549, U-87 MG), then the RNA of treated cells were used for the cDNA library synthesis and RNA sequencing. The differentially expressed genes were used for functional enrichment and the gene expression network was analyzed to find the most relevant biological process with SOX2OT function. Furthermore, the expression change of candidate genes was measured by qRT-PCR for more confirmation and the cell cycle was monitored by PI staining. RESULTS: Our findings showed that SOX2OT knockdown affects the cellular gene expression generally with enriched cell proliferation and development biological process. Particularly, the cell cycle and mitotic regulatory genes expression including: CDK2, CDK2AP2, ACTR3, and chromosome structure associated genes like SMC4, INCENP and GNL3L are changed in treated cancer cells. CONCLUSION: Our results propound SOX2OT association with cell cycle and mitosis regulation in cancer cells.
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The anti-inflammatory role of macrophages in apoptotic cells (ACs) clearance is involved in Systemic Lupus Erythematosus (SLE) pathogenesis. The efferocytic capability of macrophages is altered by M1/M2 polarization. Histone deacetylase inhibitors (HDACi) are proposed to enhance the expansion of M2 macrophages. Sodium valproate (VPA) is an HDACi with different anti-inflammatory properties. Here, we aimed to investigate the effects of HDACi by VPA on the polarization of monocyte-derived macrophages (MDMs) and regulating the expression of anti-inflammatory cytokines in SLE. We studied the ex vivo alterations of MDMs among 15 newly diagnosed SLE patients and 10 normal subjects followed by ACs and VPA treatments. M1/M2 polarization was assessed by expression of CD86/CD163, IL1-ß, IDO-1, and MRC-1 among treated and non-treated MDMs. We also evaluated the production of IL-10, IL-12, TGF-ß1, and TNF-α cytokines in the cell culture supernatants. CD163 was overexpressed upon VPA treatment, while CD86 showed no significant change. IL1-ß and IDO-1 genes were significantly downregulated, and the mRNA expression of MRC-1 was increased among VPA-treated MDMs of SLE patients. The anti-inflammatory cytokines (IL-10 and TGF-ß1) were overproduced while TNF-α level was decreased in response to VPA. The population of classically activated macrophages was more prevalent among SLE patients and efferocytosis was defected. VPA could successfully enhance the anti-inflammatory immune response through alternative activation of MDMs in SLE patients.
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Inibidores de Histona Desacetilases/farmacologia , Lúpus Eritematoso Sistêmico/imunologia , Macrófagos/efeitos dos fármacos , Ácido Valproico/farmacologia , Polaridade Celular/efeitos dos fármacos , Citocinas/metabolismo , Feminino , Humanos , Lúpus Eritematoso Sistêmico/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismoRESUMO
Despite the improvements in cancer treatment, breast cancer still remains the second most common cause of death from cancer in women. Doxorubicin (DOXO) is widely used for cancer treatment. However, drug resistance limits the treatment outcome. Here, we investigated the toxicity of DOXO in combination with an antifungal agent amphotericin B (AmB) against the MCF-7 breast cancer cell line. The cell viability was measured using MTT assay. The apoptosis was studied by caspase-8 and caspase-9 activity measurements and DNA fragmentation was investigated by TUNEL assay. The combination of two drugs significantly increased the apoptotic index and the caspase-8 and caspase-9 activities in comparison to DOXO-treated cells. Our finding showed that pre-treatment of MCF-7 cells with AmB synergistically exerted the anticancer effect of DOXO through the caspase-dependent apoptosis manner.
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Despite the advances in cancer therapy, lung cancer still remains the most leading cause of cancer death worldwide. The long non-coding RNAs (lncRNAs) are recently introduced as novel regulators of human cancers. SOX2 overlapping transcript (SOX2OT) is a cancer-associated lncRNA gene that encodes different alternatively spliced transcripts. Here, we investigated the alterations in the preferential expression of different SOX2OTs in twenty non-small cell lung cancer (NSCLC) patients by real-time quantitative reverse transcription PCR (qRT-PCR) method. We observed preferential expression of SOX2OT4 and SOX2OT7 in lung tumor tissues. The quantitative gene expression analysis revealed that >30 % of NSCLC tumors express SOX2OT4 (mean = 7.6 times) and SOX2OT7 (mean = 5.9 times) more than normal tissues, with higher expression in squamous cell carcinoma. Further, we observed overexpression of pluripotency-associated transcription factor, SOX2 in 47 % of our samples concordant with SOX2OT (R = 0.62, P value <0.05). Overexpression of OCT4A gene was also observed in 36.8 % of tumor tissues. Then, we investigated the effects of SOX2OT suppression in lung adenocarcinoma cell line, by means of RNAi. Cell characteristics of colony formation, apoptosis, 2-D mobility, and cell cycle progression were measured in control and treated A549 cells. The SOX2OT knockdown significantly reduced the colony formation ability of cancer cells; however, no alterations in the rate of apoptosis were detected. On the other hand, SOX2OT-suppressed cells had elevated accumulation in G2/M phase of cell cycle and exhibited limited mobility. Altogether, our findings support a potential oncogenic role for SOX2OT in non-small cell lung cancer tumor genesis and SOX2OT seems a promising therapeutic candidate for NSCLC.
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Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , RNA Longo não Codificante/genética , Processamento Alternativo , Feminino , Citometria de Fluxo , Variação Genética , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Transcriptoma , Regulação para CimaRESUMO
Long non-coding RNAs are manifested as a new paradigm of molecular effectors in a wide range of human diseases. Human SOX2 overlapping transcript (SOX2OT) gene can generate six lncRNA transcript variants which are functionally assumed to be correlated with cellular differentiation and carcinogenesis. However, the circumstances determining expressional and functional differences between SOX2OT transcript variants remain to be explored. Here, we studied the expression of all SOX2OT transcript variants specifically in five human cancer cell lines by real-time RT-PCR. Changes of the new SOX2OT transcript variants expression were measured during the NT2 teratocarcinoma cell line neuronal-like differentiation and were compared to pluripotency regulators, SOX2 and OCT4A gene expressions. Surprisingly, we identified two new SOX2OT transcripts, named SOX2OT-7, SOX2OT-8 which lack exon 8. We discovered that beside active proximal and distal SOX2OT promoters, different cancer cell lines express high levels of some SOX2OT transcript variants differentially by alternative splicing. Significantly, both SOX2OT-7 and SOX2OT-8 are highly expressed in human cancer cell lines coinciding with SOX2, one of the pluripotency regulators. Our results revealed that SOX2OT-7 is almost the most abundant form of SOX2OT transcript variants in the examined cancer cell lines particularly in NT2 teratocarcinoma cell line where its expression falls upon neuronal-like differentiation similar to SOX2 and OCT4A. We suggest that at least some of SOX2OT transcripts are significantly associated with cancer and stem cell related pathways.