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1.
Stem Cells Dev ; 2020 Mar 17.
Artigo em Inglês | MEDLINE | ID: mdl-32178572

RESUMO

Laryngeal squamous cell carcinoma (LSCC) is a common head and neck cancer that is unresponsive to chemotherapy; therefore, understanding the causes of chemotherapy resistance is important. The cancer stem cell theory postulates that cancer stem cells (CSCs) are the source of tumor chemoresistance. We enrich laryngeal CSCs to overcome chemoresistance of LSCC. A laryngeal cancer xenograft model was established, and a low dose of cisplatin was administered until chemoresistance arose. A next-generation xenograft model was established using surviving tumor cells, and the test was repeated 4 times to screen for CSCs. Cell-function experiments were performed on each tumor cell generation (m1, m2, m3, and m4). The m3 line, with the highest stemness, was selected for transcriptome sequencing. LY6D was selected for clinical sample validation and functional verification. LY6D expression was detected in 107 laryngeal cancer samples, with high expression in 91 of these samples. LY6D expression was correlated with pathological T- and clinical stages, and with cervical lymph node metastasis. The siLY6D group exhibited reduced adhesion and chemoresistance to cisplatin, 5-fluorouracil, and paclitaxel. LY6D is upregulated in laryngeal cancer and may serve as a biomarker for chemoresistance in CSCs. Moreover, LY6D could serve as an alternative antigenic peptide in the targeted treatment of laryngeal cancer.

2.
Mol Cell Biol ; 2020 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-31988106

RESUMO

Intracellular pathogen resistance 1 (Ipr1) is found to be a mediator to integrate cGMP-cAMP synthase (cGAS) - interferon regulatory factor 3 (IRF3), activated by intracellular pathogens, with p53 pathway. Previous studies have shown the process of Ipr1 induction by various immune reactions, including intracellular bacterial and viral infection. The present study identified that Ipr1 was regulated by cGAS-IRF3 pathway during pathogenic infection. IRF3 was found to regulate Ipr1 expression by directly binding the interferon-stimulated response element motif of the Ipr1 promoter. Knockdown of Ipr1 decreased the expression of immunity-related GTPase family M member 1 (Irgm1) which plays critical roles in autophagy initiation. Irgm1 promoter characterization revealed a p53 motif in front of the transcription start site. P53 was found to participate in regulation of Irgm1 expression and Ipr1-related effects on p53 stability by affecting interactions between ribosomal protein L11 (Rpl11) and transformed mouse 3T3 cell double minute 2 (Mdm2). Our results indicate Ipr1 integrates cGAS-IRF3 with p53 modulated Irgm1 expression.

3.
Onco Targets Ther ; 12: 10441-10453, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31819525

RESUMO

Background: Recent studies revealed that miR-424-5p regulates the malignant behavior of multiple cancer types. However, the expression and function of miR-424-5p in laryngeal squamous cell carcinoma (LSCC) is unclear. Purpose: This study aimed to evaluate the association of miR-424-5p level with clinical features of LSCC and investigate the effect and potential mechanism of miR-424-5p on LSCC progression. Methods: The expression of miR-424-5p in LSCC and paired adjacent normal margin (ANM) tissues from 106 patients with LSCC were analyzed by quantitative PCR (qPCR), and clinical significance was analyzed. Target genes of miR-424-5p were predicted, followed by functional annotation. The functional role of miR-424-5p in LSCC was investigated by molecular and cellular experiments with LSCC cell lines, with flow cytometry used for cell cycle analysis. In addition, miR-424-5p regulation of the predicted target gene cell adhesion molecule 1 (CADM1) was validated by qPCR, Western blot analysis and luciferase reporter assay. Results: miR-424-5p was upregulated in LSCC versus ANM tissues. High miR-424-5p level was significantly associated with poor differentiation, advanced tumor stage and cervical lymph node metastasis. Bioinformatics analysis showed that miR-424-5p target genes are mainly enriched in biological processes of the cell cycle, cell division, and negative regulation of cell migration, and were involved in multiple cancer-related pathways. Overexpression of miR-424-5p promoted proliferation, migration, invasion, and adhesion of LSCC cells and affected the cell cycle progression. Additionally, CADM1 was a direct target of miR-424-5p in LSCC cells. Conclusion: miR-424-5p functions as an oncogene to promote the aggressive progression of LSCC, and CADM1 is a direct downstream target of miR-424-5p in LSCC cells. miR-424-5p may be a potential therapeutic target in LSCC.

4.
Nat Commun ; 10(1): 5334, 2019 11 25.
Artigo em Inglês | MEDLINE | ID: mdl-31767869

RESUMO

Protein products of the regenerating islet-derived (REG) gene family are important regulators of many cellular processes. Here we functionally characterise a non-protein coding product of the family, the long noncoding RNA (lncRNA) REG1CP that is transcribed from a DNA fragment at the family locus previously thought to be a pseudogene. REG1CP forms an RNA-DNA triplex with a homopurine stretch at the distal promoter of the REG3A gene, through which the DNA helicase FANCJ is tethered to the core promoter of REG3A where it unwinds double stranded DNA and facilitates a permissive state for glucocorticoid receptor α (GRα)-mediated REG3A transcription. As such, REG1CP promotes cancer cell proliferation and tumorigenicity and its upregulation is associated with poor outcome of patients. REG1CP is also transcriptionally inducible by GRα, indicative of feedforward regulation. These results reveal the function and regulation of REG1CP and suggest that REG1CP may constitute a target for cancer treatment.

5.
J Cancer ; 10(19): 4455-4462, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31528209

RESUMO

Laryngeal cancer has the second highest incidence of head and neck malignant tumors worldwide. In recent years, studies have shown that human papillomavirus (HPV) infection may be a high-risk factor for laryngeal cancer and closely related to the development and prognosis of laryngeal cancer. The mechanism of the occurrence and development of laryngeal cancer caused by HPV infection needs investigation, as does a rapid and effective HPV detection method for effectively preventing the occurrence of laryngeal cancer and controlling its development. Many studies have explored the relation between HPV infection and laryngeal cancer. Here we review the research progress in investigating HPV infection in terms of DNA, mRNA and protein levels in the occurrence and development of laryngeal cancer and routine HPV detection methods.

6.
IUBMB Life ; 71(11): 1771-1784, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31298480

RESUMO

Fascin actin-bundling protein 1 (FSCN1) is an evolutionarily conserved actin-bundling protein that plays a critical role in cell migration, motility, adhesion, and cellular interactions. Although multiple clinical studies have implicated the expression of FSCN1 in laryngeal squamous cell carcinoma (LSCC) progression, the precise mechanism of FSCN1 in the process has not been clearly elucidated. To define FSCN1 function, we characterized FSCN1­interacting proteins in two cell lines by immunoprecipitation followed by mass spectrometry (MS). After data filtering, 119 proteins with expression in both the Hep-2 and TU-177 cell samples were identified as FSCN1-interacting partners. With in-depth bioinformatics analysis, we linked FSCN1 to critical cellular processes including cell adhesion, glycolysis/gluconeogenesis, regulation of protein ubiquitination, ribosomal RNA processing, and small molecule metabolism. We discuss the interactions between FSCN1 and some of the newly validated partners. The identification of these potential partners of FSCN1 expands our knowledge of the FSCN1 interactome and provides a valuable resource for understanding the functions of this protein in LSCC progression.

7.
Proteomics ; 19(21-22): e1900059, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31287215

RESUMO

Dysregulation of fascin actin-bundling protein 1 (FSCN1) enhances cell proliferation, invasion, and motility in laryngeal squamous cell carcinoma (LSCC), while the mechanism remains unclear. Here, co-immunoprecipitation and mass spectrometry is utilized to identify potential FSCN1-binding proteins. Functional annotation of FSCN1-binding proteins are performed by Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis. Furthermore, the protein-protein interaction network of FSNC1-binding proteins is constructed and the interactions between FSCN1 and novel identified interacting proteins AIMP1 and LTA4H are validated. Moreover, the expression and functional role of AIMP1 and LTA4H in LSCC are investigated. A total of 123 proteins are identified as potential FSCN1-binding proteins, and functional annotation shows that FSCN1-binding proteins are significantly enriched in carcinogenic processes, such as filopodium assembly-regulation and GTPase activity. Co-IP/western blotting and immunofluorescence confirm that AIMP1 and LTA4H bind and colocalize with FSCN1. Furthermore, both AIMP1 and LTA4H are upregulated in LSCC tissues, and knockdown of AIMP1 or LTA4H inhibits LSCC cell proliferation, migration, and invasion. Collectively, the identification of FSCN1-binding partners enhances understanding of the mechanism of FSCN1-mediated malignant phenotypes, and these findings indicate that FSCN1 binds to AIMP1 and LTA4H might promote the progression of LSCC.

8.
Proteomics ; 19(21-22): e1900020, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31169343

RESUMO

This study intends to investigate the transcriptional regulatory role of miR-145-5p in laryngeal squamous cell carcinoma (LSCC). LSCC cell line TU-177 is transfected with miR-145-5p mimics, generating miR-145-5p-overexpression LSCC cells. Whole transcriptome microarrays are used to investigate the differentially expressed lncRNAs, circRNAs, mRNAs, and miRNAs. The target genes of miRNAs are predicted and performed functional annotation. Additionally, the circRNAs, lncRNAs, and mRNAs that interact with miRNAs are predicted, and then the competing endogenous RNAs (ceRNAs) are predicted. Microarray analysis identifies 26 miRNAs, 248 mRNAs, 1118 lncRNAs, and 382 circRNAs differentially expressed in miR-145-5p overexpressed LSCC cells. Overall, 675 target genes are identified for the differentially expressed miRNAs, which involved in cell adhesion associated gene ontology (GO) terms, and MAPK and FoxO signaling pathways. The up-regulated mRNAs involved in the pathway of ABC transporters, while the down-regulated mRNAs involved in pathway of olfactory transduction. Moreover, 149 ceRNAs are predicted, which are associated with apoptosis, Wnt pathway, and metabolic pathway. Furthermore, qPCR results confirm that miR-145-5p affects expression of lncRNAs, miRNAs, mRNAs, and circRNAs in LSCC cells. Collectively, miR-145-5p may be inhibits LSCC progression via ceRNA-mediated pathways, such as WNT2B-miR-145-5p-NONHSAT127539.2, CASP10-miR-145-5p-NONHSAT127539.2, CASP10-miR-145-5p-circ_0003519, and TPO-miR-145-5p-circ_0003519.

9.
Nucleic Acids Res ; 47(W1): W315-W321, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31069394

RESUMO

Identifying the variants that alter protein function is a promising strategy for deciphering the biological consequences of somatic mutations during tumorigenesis, which could provide novel targets for the development of cancer therapies. Here, based on our previously developed method, we present a strategy called AlloDriver that identifies cancer driver genes/proteins as possible targets from mutations. AlloDriver utilizes structural and dynamic features to prioritize potentially functional genes/proteins in individual cancers via mapping mutations generated from clinical cancer samples to allosteric/orthosteric sites derived from three-dimensional protein structures. This strategy exhibits desirable performance in the reemergence of known cancer driver mutations and genes/proteins from clinical samples. Significantly, the practicability of AlloDriver to discover novel cancer driver proteins in head and neck squamous cell carcinoma (HNSC) was tested in a real case of human protein tyrosine phosphatase, receptor type K (PTPRK) through a L1143F driver mutation located at the allosteric site of PTPRK, which was experimentally validated by cell proliferation assay. AlloDriver is expected to help to uncover innovative molecular mechanisms of tumorigenesis by perturbing proteins and to discover novel targets based on cancer driver mutations. The AlloDriver is freely available to all users at http://mdl.shsmu.edu.cn/ALD.

10.
Mol Ther ; 27(2): 365-379, 2019 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-30341010

RESUMO

Laryngeal squamous cell carcinoma (LSCC) is a common form of head and neck cancer with poor prognosis. However, the mechanism underlying the pathogenesis of LSCC remains unclear. Here, we demonstrated increased expression of fascin actin-bundling protein 1 (FSCN1) and decreased expression of microRNA-145-5p (miR-145-5p) in a clinical cohort of LSCC. Luciferase assay revealed that miR-145-5p is a negative regulator of FSCN1. Importantly, low miR-145-5p expression was correlated with TNM (tumor, node, metastasis) status and metastasis. Moreover, cases with low miR-145-5p/high FSCN1 expression showed poor prognosis, and these characteristics together served as independent prognostic indicators of survival. Gain- and loss-of-function studies showed that miR-145-5p overexpression or FSCN1 knockdown inhibited LSCC migration, invasion, and growth by suppressing the epithelial-mesenchymal transition along with inducing cell-cycle arrest and apoptosis. Additionally, hypermethylation of the miR-145-5p promoter suggested that repression of miR-145-5p arises through epigenetic inactivation. LSCC tumor growth in vivo could be inhibited by using miR-145-5p agomir or FSCN1 small interfering RNA (siRNA), which highlights the potential for clinical translation. Collectively, our findings indicate that miR-145-5p plays critical roles in inhibiting the progression of LSCC by suppressing FSCN1. Both miR-145-5p and FSCN1 are important potential prognostic markers and therapeutic targets for LSCC.


Assuntos
Proteínas de Transporte/metabolismo , Metilação de DNA/fisiologia , Neoplasias de Cabeça e Pescoço/genética , Neoplasias de Cabeça e Pescoço/metabolismo , Neoplasias Laríngeas/genética , Neoplasias Laríngeas/metabolismo , MicroRNAs/genética , Proteínas dos Microfilamentos/metabolismo , Regiões Promotoras Genéticas/genética , Animais , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Proteínas de Transporte/genética , Linhagem Celular , Linhagem Celular Tumoral , Metilação de DNA/genética , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/fisiologia , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/fisiologia , Proteínas dos Microfilamentos/genética
11.
Cell Physiol Biochem ; 47(4): 1696-1710, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29949786

RESUMO

BACKGROUND/AIMS: CD133+CD44+ cancer stem cells previously isolated from laryngeal squamous cell carcinoma (LSCC) cell lines showed strong malignancy and tumorigenicity. However, the molecular mechanism underlying the enhanced malignancy remained unclear. METHODS: Cell proliferation assay, spheroid-formation experiment, RNA sequencing (RNA-seq), miRNA-seq, bioinformatic analysis, quantitative real-time PCR, migration assay, invasion assay, and luciferase reporter assay were used to identify differentially expressed mRNAs, lncRNAs, circRNAs and miRNAs, construct transcription regulatory network, and investigate functional roles and mechanism of circRNA in CD133+CD44+ laryngeal cancer stem cells. RESULTS: Differentially expressed genes in TDP cells were mainly enriched in the biological processes of cell differentiation, regulation of autophagy, negative regulation of cell death, regulation of cell growth, response to hypoxia, telomere maintenance, cellular response to gamma radiation, and regulation of apoptotic signaling, which are closely related to the malignant features of tumor cells. We constructed the regulatory network of differentially expressed circRNAs, miRNAs and mRNAs. qPCR findings for the expression of key genes in the network were consistent with the sequencing data. Moreover, our data revealed that circRNA hg19_circ_0005033 promotes proliferation, migration, invasion, and chemotherapy resistance of laryngeal cancer stem cells. CONCLUSIONS: This study provides potential biomarkers and targets for LSCC diagnosis and therapy, and provide important evidences for the heterogeneity of LSCC cells at the transcription level.


Assuntos
Antígeno AC133/metabolismo , Antígenos CD/metabolismo , Biomarcadores Tumorais/metabolismo , Caderinas/metabolismo , Carcinoma de Células Escamosas/metabolismo , Perfilação da Expressão Gênica , Neoplasias Laríngeas/metabolismo , Proteínas de Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Humanos , Células-Tronco Neoplásicas/patologia
12.
J Immunol ; 200(10): 3506-3518, 2018 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-29661829

RESUMO

Mycobacterium tuberculosis poses a significant global health threat. MicroRNAs play an important role in regulating host anti-mycobacterial defense; however, their role in apoptosis-mediated mycobacterial elimination and inflammatory response remains unclear. In this study, we explored the role of microRNA-27b (miR-27b) in murine macrophage responses to M. tuberculosis infection. We uncovered that the TLR-2/MyD88/NF-κB signaling pathway induced the expression of miR-27b and miR-27b suppressed the production of proinflammatory factors and the activity of NF-κB, thereby avoiding an excessive inflammation during M. tuberculosis infection. Luciferase reporter assay and Western blotting showed that miR-27b directly targeted Bcl-2-associated athanogene 2 (Bag2) in macrophages. Overexpression of Bag2 reversed miR-27b-mediated inhibition of the production of proinflammatory factors. In addition, miR-27b increased p53-dependent cell apoptosis and the production of reactive oxygen species and decreased the bacterial burden. We also showed that Bag2 interacts with p53 and negatively regulates its activity, thereby controlling cell apoptosis and facilitating bacterial survival. In summary, we revealed a novel role of the miR-27b/Bag2 axis in the regulation of inflammatory response and apoptosis and provide a potential molecular host defense mechanism against mycobacteria.


Assuntos
Apoptose/genética , Inflamação/genética , MicroRNAs/genética , Tuberculose/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Animais , Linhagem Celular , Feminino , Células HEK293 , Humanos , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/patogenicidade , NF-kappa B/genética , Proteínas Proto-Oncogênicas c-bcl-2/genética , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/genética , Tuberculose/metabolismo
13.
Cell Physiol Biochem ; 44(5): 2057-2072, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29241165

RESUMO

BACKGROUND/AIMS: Self-renewal is one of the most important features of embryonic stem (ES) cells. SC1 is a small molecule modulator that effectively maintains the self-renewal of mouse ES cells in the absence of leukemia inhibitory factor (LIF), serum and feeder cells. However, the mechanism by which SC1 maintains the undifferentiated state of mouse ES cells remains unclear. METHODS: In this study, microarray and small RNA deep-sequencing experiments were performed on mouse ES cells treated with or without SC1 to identify the key genes and microRNAs that contributed to self-renewal. RESULTS: SC1 regulates the expressions of pluripotency and differentiation factors, and antagonizes the retinoic acid (RA)-induced differentiation in the presence or absence of LIF. SC1 inhibits the MEK/ERK pathway through Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis and pathway reporting experiments. Small RNA deep-sequencing revealed that SC1 significantly modulates the expression of multiple microRNAs with crucial functions in ES cells. The expression of miR124-3p is upregulated in SC1-treated ES cells, which significantly inhibits the MEK/ERK pathway by targeting Grb2, Sos2 and Egr1. CONCLUSION: SC1 enhances the self-renewal capacity of mouse ES cells by modulating the expression of key regulatory genes and pluripotency-associated microRNAs. SC1 significantly upregulates miR124-3p expression to further inhibit the MEK/ ERK pathway by targeting Grb2, Sos2 and Egr1.


Assuntos
Autorrenovação Celular/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , MicroRNAs/metabolismo , Células-Tronco Embrionárias Murinas/citologia , Pirazóis/farmacologia , Pirimidinas/farmacologia , Animais , Diferenciação Celular/efeitos dos fármacos , Proteína 1 de Resposta de Crescimento Precoce/antagonistas & inibidores , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Proteína Adaptadora GRB2/antagonistas & inibidores , Proteína Adaptadora GRB2/metabolismo , Fator Inibidor de Leucemia/química , MAP Quinase Quinase Quinases/metabolismo , Camundongos , MicroRNAs/química , MicroRNAs/genética , Células-Tronco Embrionárias Murinas/metabolismo , Análise de Sequência de RNA , Proteínas Son Of Sevenless/antagonistas & inibidores , Proteínas Son Of Sevenless/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Tretinoína/farmacologia , Regulação para Cima/efeitos dos fármacos
14.
Oncotarget ; 8(38): 64050-64065, 2017 Sep 08.
Artigo em Inglês | MEDLINE | ID: mdl-28969051

RESUMO

Tuberculosis remains a leading health problem worldwide and still accounts for about 1.3 million deaths annually. Expression of the mouse Sp110 nuclear body protein (Sp110) upregulates the apoptotic pathway, which plays an essential role in enhancing host immunity to Mycobacterium tuberculosis (Mtb). However, the mechanism of this upregulation is unclear. Here, we have identified 253 proteins in mouse macrophages that interact with Sp110, of which 251 proteins were previously uncharacterized. The results showed that Sp110 interacts with heat shock protein 5 (Hspa5) to activate endoplasmic reticulum (ER) stress-induced apoptosis, and that this is essential for Sp110 enhanced macrophage resistance to Mtb. Inhibition of the ER stress pathway abolishing the Sp110-enhanced macrophage apoptosis and resulted in increased intracellular survival of Mtb in macrophages overexpressing Sp110 Further studies revealed that Sp110 also interacts with the RNA binding protein, Ncl to promote its degradation. Consequently, the expression of Bcl2, usually stabilized by Ncl, was downregulated in Sp110 overexpressing macrophages. Moreover, overexpression of Sp110 promotes degradation of ribosomal protein Rps3a, resulting in upregulation of the activity of the pro-apoptotic poly (ADP-ribose) polymerase (PARP). In addition, macrophages from transgenic cattle with increased Sp110 expression confirmed that activation of the ER stress response is the main pathway through which Sp110-enhanced macrophages impart resistance to Mtb. This work has revealed the mechanism of Sp110 enhanced macrophage apoptosis in response to Mtb infection, and provides new insights into the study of host-pathogen interactions.

15.
J Cancer ; 8(12): 2356-2368, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28819440

RESUMO

Background and aim: Understanding the molecular biological mechanisms underlying laryngeal squamous cell carcinoma (LSCC) invasion and metastasis is crucial for diagnosis, treatment, and prognosis. We aimed to examine the expression of the tumor suppressor microRNA-204-5p (miR-204-5p) and its target gene, forkhead box C1 (FOXC1), in human LSCC and explore their roles in the malignant behaviors of LSCC Hep-2 and TU-177 cells. Methods: The regulatory effects of miR-204-5p on the 3' untranslated region of FOXC1 predicted by bioinformatics were tested by dual-luciferase reporter assay. Quantitative RT-PCR was used to detect mRNA expression in 43 fresh samples of LSCC and corresponding adjacent normal mucosa (ANM). FOXC1 protein expression was examined by immunohistochemistry. miR-204-5p mimics and FOXC1 siRNA were transfected into LSCC cell lines Hep-2 and TU-177 to observe malignant behavior. miR-204-5p mimics were injected into Hep-2 or TU-177 xenograft tumors in nude mice to examine tumor growth.Results: The miR-204-5p mRNA level was lower in all 43 LSCC samples than in the ANM samples, but the FOXC1 level was higher in the LSCC samples than in the ANM samples. The miR-204-5p level was lower for stage III and IV cancer and lymph node N+ status samples than for stage I and II cancer and N0 status samples. FOXC1 mRNA and protein levels were higher for N+ than for N0 LSCC. The miR-204-5p mRNA levels were lower in Hep-2 and TU-177 cells than in ANM tissues, but FOXC1 mRNA levels were higher in Hep-2 and TU-177 cells than in ANM tissues. Dual-luciferase reporter assays demonstrated the targeted regulatory effects of miR-204-5p on the FOXC1 3' UTR. Cell proliferation and colony formation was facilitated with miR-204-5p mimics and FOXC1 siRNA, with weaker cell migration and invasion than the controls. Moreover, miR-204-5p overexpression or FOXC1 knockdown inhibited the EMT process in LSCC cells. In vivo experiments demonstrated that injection of miR-204-5p into Hep-2 and TU-177 xenograft tumors in nude mice significantly inhibited tumor growth. Conclusions: miR-204-5p is involved in the invasion and metastasis of LSCC. It has a targeted regulatory effect on FOXC1 expression; malignant LSCC behaviors, including cell proliferation, invasion, and metastasis, are suppressed, and tumor growth in vivo is inhibited. This suggests that miR-204-5p may be a target for molecular therapy of LSCC in the future.

16.
Mol Med Rep ; 16(5): 5863-5870, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28849105

RESUMO

The present study aimed to investigate the variations of the gene network and biological functions induced by hsa­miR­145­5p in the laryngeal squamous cell carcinoma (LSCC) cell line Tu­177. A hsa­miR­145­5p­overexpressed Tu­177 cell model was established, and the gene expression microarray data of miR­145­5p­overexpressed cells and negative control (NC) cells were analyzed. The differentially expressed genes (DEGs) between two groups were identified, and their potential functions were predicted by functional enrichment analysis. Furthermore, the targets of miR­145­5p were identified from the DEGs, and their potential functions and protein­protein interactions (PPIs) were analyzed. The mRNA expressions of acetyl­CoA carboxylase ß (ACACB), fibroblast growth factor receptor 1 (FGFR1), protein phosphatase 3 catalytic subunit a (PPP3CA) and spleen associated tyrosine kinase (SYK), were analyzed via quantitative polymerase chain reaction. A total of 1,501 upregulated and 887 downregulated genes were identified in the hsa­miR­145­5p­overexpressed Tu­177 cells, compared with the NC cells. Of these DEGs, 164 upregulated and 221 downregulated genes were predicted to be targeted by hsa­miR­145­5p. The upregulated target genes were primarily associated with functions of immunity, whereas the downregulated target genes were significantly enriched in the p53 signaling pathway. In the PPI network consisting of 267 target genes, the upregulated ACACB had the greatest degree and interacted with downregulated genes including PPP3CA and SYK, in addition to upregulated genes, including FGFR1. The mRNA expressions of ACACB and FGFR1were markedly enhanced in miR­145­5p­overexpressed Tu­177 cells, whereas overexpressing miR­145­5p significantly reduced mRNA expression of PPP3CA and SYK. hsa­miR­145­5p may exhibit an anticancer role in LSCC via regulating multiple cell processes, including cell proliferation and invasion, fatty acid metabolism, immunity and p53 signaling pathway. These findings provide novel information for the future investigation of miR­145­5p functions in LSCC.


Assuntos
Carcinoma de Células Escamosas/genética , Neoplasias Laríngeas/genética , MicroRNAs/genética , Mapas de Interação de Proteínas/genética , Carcinoma de Células Escamosas/patologia , Linhagem Celular Tumoral , Biologia Computacional , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Laríngeas/patologia , Análise em Microsséries , Transdução de Sinais/genética
17.
J Cancer ; 8(3): 497-506, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28261352

RESUMO

Background: Laryngeal squamous cell carcinoma ranks second among head and neck squamous-cell carcinomas. Cancer stem cells can support cancer growth and malignant behavior. Therefore, cancer stem cells isolated from laryngeal squamous cell carcinoma tissue could be used to investigate the initiation, progression, and treatment strategies of this cancer. Methods: We isolated CD133-CD44-, CD133-CD44+, CD133+CD44- and CD133+CD44+ cell populations from laryngeal squamous-cell carcinoma cell lines Hep2 and TU-177 by magnetic-activated cell sorting. Sphere formation, cell proliferation, migration, invasion, colony formation, resistance to radio- and chemotherapy, and in vivo tumorigenicity of these populations were evaluated. Moreover, we investigated the expression of the stem-cell markers (sex determining region Y)-box 2 (SOX2) and octamer-binding transcription factor 4 (OCT4) in CD133-CD44-, CD133-CD44+, CD133+CD44-, CD133+CD44+ cell populations and parental Hep2 and TU-177 cells. Results: As compared with CD133-CD44-, CD133-CD44+, CD133+CD44- populations and parental cells, CD133+CD44+ cells showed higher cell viability, migration and invasive capability and colony formation ability as well as stronger resistance to cisplatin and irradiation. Moreover, levels of SOX2 and OCT4 and tumorigenicity in nude mice were greater in CD133+CD44+ Hep2 and TU-177 cells than other cell populations and parental cells. Conclusion: The CD133+CD44+ population of laryngeal squamous-cell carcinoma Hep2 and TU-177 cells have stem cell properties and showed more malignant features than CD133+CD44- and CD133-CD44+ cell populations. CD133+CD44+ cancer stem cells may be a promising target for developing anticancer drugs and treatment strategies for laryngeal squamous cell carcinoma.

18.
J Cell Sci ; 130(10): 1740-1751, 2017 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-28348106

RESUMO

In murine macrophages infected with Mycobacterium tuberculosis (Mtb), the level of phosphorylated STAT1 (P-STAT1), which drives the expression of many pro-apoptosis genes, increases quickly but then declines over a period of hours. By contrast, infection induces a continued increase in the level of unphosphorylated STAT1 that persists for several days. Here, we found that the level of unphosphorylated STAT1 correlated with the intracellular bacterial burden during the later stages of infection. To investigate the significance of a high level of unphosphorylated STAT1, we increased its concentration exogenously, and found that the apoptosis rate induced by Mtb was sufficiently decreased. Further experiments confirmed that unphosphorylated STAT1 affects the expression of several immune-associated genes and lessens the sensitivity of macrophages to CD95 (FAS)-mediated apoptosis during Mtb infection. Furthermore, we characterized 149 proteins that interacted with unphosphorylated STAT1 and the interactome network. The cooperation between unphosphorylated STAT1 and STAT3 results in downregulation of CD95 expression. Additionally, we verified that unphosphorylated STAT1 and IFIT1 competed for binding to eEF1A. Taken together, our data show that the role of unphosphorylated STAT1 differs from that of P-STAT1, and represses apoptosis in macrophages to promote immune evasion during Mtb infection.


Assuntos
Apoptose , Macrófagos/metabolismo , Macrófagos/microbiologia , Mycobacterium tuberculosis/fisiologia , Fator de Transcrição STAT1/metabolismo , Animais , Ligação Competitiva , Proteínas de Transporte/metabolismo , Proteína Ligante Fas/metabolismo , Macrófagos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Fator 1 de Elongação de Peptídeos/metabolismo , Fosforilação , Mapas de Interação de Proteínas , Células RAW 264.7 , Proteínas de Ligação a RNA , Fator de Transcrição STAT3/metabolismo , Transcrição Genética , Tuberculose/metabolismo , Tuberculose/microbiologia , Receptor fas/genética , Receptor fas/metabolismo
19.
Theriogenology ; 92: 156-166, 2017 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-28237332

RESUMO

Peroxiredoxin 5 (PRDX5) is an important peroxiredoxin with antioxidant and antiapoptotic functions. The objective of this study was to determine whether the antioxidant and antiapoptotic activities of PRDX5 play an essential role in the in vitro development of bovine SCNT embryos. We analyzed the expression patterns of PRDX5, and found the expression of PRDX5 in second meiotic metaphase oocytes, which persisted until the blastocyst stage IVF and SCNT embryos. In addition, PRDX5 transcript levels were significantly higher in SCNT embryos than those in IVF embryos. After PRDX5 inhibition, more embryos were arrested before eight-cell stage (53.0% vs. 35.4%, P < 0.05) and decreases were observed in the blastocyst rate (15.7% vs. 32.3%, P < 0.05), inner cell mass rate (24.45 ± 6.05% vs. 38.37 ± 6.83%, P < 0.05), and total cell number (90.8 ± 10.2 vs. 116.2 ± 10.8, P < 0.05). There were significant increases in reactive oxygen species levels at two-, four-, and eight-cell stages and the apoptosis rate of blastocysts (11.93 ± 2.12% vs. 4.25 ± 0.88%, P < 0.05). The expression levels of p53 and HSP70 were increased significantly, whereas the expression level of Bcl-2 was decreased significantly. These results suggest that the antioxidant and antiapoptotic activities of PRDX5 play important roles in the in vitro development of bovine SCNT embryos.


Assuntos
Bovinos/embriologia , Técnicas de Cultura Embrionária/veterinária , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Técnicas de Transferência Nuclear/veterinária , Peroxirredoxinas/metabolismo , Animais , Anticorpos Monoclonais , Blastocisto/fisiologia , Peroxirredoxinas/genética
20.
PLoS One ; 11(9): e0162832, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-27622275

RESUMO

The mouse intracellular pathogen resistance 1 (Ipr1) gene plays important roles in mediating host immunity and previous work showed that it enhances macrophage apoptosis upon mycobacterium infection. However, to date, little is known about the regulation pattern of Ipr1 action. Recent studies have investigated the protein-coding genes and microRNAs regulated by Ipr1 in mouse macrophages, but the structure and the functional motif of the Ipr1 protein have yet to be explored. In this study, we analyzed the domains and functional motif of the Ipr1 protein. The resulting data reveal that Ipr1 protein forms a homodimer and that the Sp100-like domain mediates the targeting of Ipr1 protein to nuclear dots (NDs). Moreover, we found that an Ipr1 mutant lacking the classic nuclear localization signal (cNLS) also translocated into the nuclei, suggesting that the cNLS is not the only factor that directs Ipr1 nuclear localization. Additionally, mechanistic studies revealed that an arginine/lysine-rich element within the DNA-binding domain (SAND domain) is critical for Ipr1 binding to the importin protein receptor NPI-1, demonstrating that this element plays an essential role in mediating the nuclear localization of Ipr1 protein. Furthermore, our results show that this arginine/lysine-rich element contributes to the transcriptional regulation and apoptotic activity of Ipr1. These findings highlight the structural foundations of Ipr1 action and provide new insights into the mechanism of Ipr1-mediated resistance to mycobacterium.


Assuntos
Transativadores/química , Transativadores/metabolismo , Sequência de Aminoácidos , Animais , Arginina/química , Núcleo Celular/metabolismo , DNA/metabolismo , Células HEK293 , Humanos , Lisina/química , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Mycobacterium tuberculosis/patogenicidade , Células NIH 3T3 , Sinais de Localização Nuclear/química , Sinais de Localização Nuclear/genética , Sinais de Localização Nuclear/metabolismo , Domínios Proteicos , Multimerização Proteica , Células RAW 264.7 , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Homologia de Sequência de Aminoácidos , Transativadores/genética , alfa Carioferinas/metabolismo
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