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Cervical cancer is the fourth most common malignant tumors in female worldwide. Cirular RNAs (circRNA) represent a new class of regulatory RNA and play a pivotal role in the carcinogenesis and development of tumors. However, their functions have not been fully elucidated in cervical cancer. In this study, we identified an upregulated circRNA, circ_0001589, both in fresh clinical samples and tissue microarray of cervical cancer. Transwell assay and cell apoptosis assay by flow cytometry demonstrated circ_0001589 promotes epithelial-mesenchymal transition (EMT)-mediated cell migration and invasion, and enhanced cisplatin resistance in vitro. In addition, in nude mice model, circ_0001589 increased the number of lung metastases and recovered xenograft growth from cisplatin treatment in vivo. Mechanistically, RNA pull-down assay, RNA immunoprecipitation, and dual-luciferase reporter assay disclosed that circ_0001589 function as an competing endogenous RNA to sponge miR-1248, which directly target the 3' untranslated region of high mobility group box-B1 (HMGB1). Thereby, circ_0001589 upregulated HMGB1 protein expression and accelerate cervical cancer progression. The rescue experiments also revealed that miR-1248 overexpression or HMGB1 knockdown partially reversed the regulatory functions of circ_0001589 on cell migration, invasion, and cisplatin resistance. In summary, our findings suggest the upregulation of circ_0001589 promoted EMT-mediated cell migration and invasion, and enhanced cisplatin resistance via regulating miR-1248/HMGB1 axis in cervical cancer. These results provided new evidence for understanding the carcinogenesis mechanism and finding new therapeutic target for cervical cancer.
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Proteína HMGB1 , MicroRNAs , Neoplasias Retais , Neoplasias do Colo do Útero , Animais , Feminino , Humanos , Camundongos , Regiões 3' não Traduzidas , Carcinogênese , Linhagem Celular Tumoral , Proliferação de Células , Cisplatino/farmacologia , Transição Epitelial-Mesenquimal , Camundongos Nus , MicroRNAs/genética , RNA Circular/genética , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/genéticaRESUMO
Increased circulating syncytiotrophoblast microparticles (STBMs) are often associated with preeclampsia (PE) but the molecular mechanisms regulating STBM shedding remain elusive. Experimental evidence has shown that actin plays a key role in STBM shedding and that Rho/ROCK is important in regulating actin rearrangement. To investigate the role of RhoB/ROCK-regulated actin arrangement in STBM shedding in PE, chorionic villous explants were prepared from placenta of patients with normotensive or PE pregnancies and BeWo cells were fused to imitate syncytiotrophoblasts. The oxygen-glucose deprivation (OGD) conditions were applied to imitate the pathophysiology of PE in vitro. The results showed that RhoB and ROCK were activated in the preeclamptic placenta, accompanied by increased actin polymerization and decreased outgrowing microvilli. In villous tissue cultures or BeWo cells, OGD activated RhoB, ROCK1 and ROCK2 and promoted STBM shedding and actin stress fibers formation. In BeWo cells, RhoB overexpression activated ROCK1 and ROCK2, leading to F-actin redistribution and STBM shedding and the OGD-induced actin polymerization and STBM shedding could be reversed by RhoB or ROCK knockdown. These results reveal that RhoB and ROCK play a key role in PE by targeting STBM shedding through actin rearrangement and that RhoB/ROCK intervention may be a potential therapeutic strategy for PE.
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Micropartículas Derivadas de Células/metabolismo , Glucose/deficiência , Oxigênio/farmacologia , Pré-Eclâmpsia/metabolismo , Pré-Eclâmpsia/patologia , Trofoblastos/metabolismo , Quinases Associadas a rho/metabolismo , Proteína rhoB de Ligação ao GTP/metabolismo , Actinas/metabolismo , Apoptose , Linhagem Celular Tumoral , Ativação Enzimática , Feminino , Humanos , Microvilosidades/metabolismo , Polimerização , GravidezRESUMO
The aberrant activation of telomerase is critical for the initiation and development of human cervical cancer, which is dependent on the activation of human telomerase reverse transcriptase (hTERT). Recently, Pin2/TRF1-interacting protein X1 (PinX1) has been identified as a suppressor of hTERT. It has been found that the telomerase is activated while the level of PinX1 is decreased in cervical cancer. However, the regulatory mechanism of PinX1 in cervical cancer cells remains unclear. In the present study, we demonstrated that the level of PinX1 is regulated by p53, and p53 functions as a transcriptional factor to directly activate the expression of PinX1 in cervical cancer cells. Moreover, we found that HPV16 E6 suppresses the expression of PinX1 via inhibiting p53 transcriptional activity, resulting in the enhancement of telomerase activity. This study not only for the first time shows that PinX1 is a novel target gene of p53 but also suggests that suppression of p53/PinX1 pathway may be a novel mechanism by which HPV16 E6 enhances the telomerase activity in cervical cancer cells.
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Proteínas Oncogênicas Virais/genética , Proteínas Repressoras/genética , Proteína Supressora de Tumor p53/genética , Proteínas Supressoras de Tumor/genética , Western Blotting , Proteínas de Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Mutação , Proteínas Oncogênicas Virais/metabolismo , Regiões Promotoras Genéticas/genética , Ligação Proteica , Interferência de RNA , Proteínas Repressoras/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Telomerase/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologiaRESUMO
AIMS: Previous studies have revealed that the increased shedding of syncytiotrophoblast extracellular vesicles (STBM) may lead to preeclampsia (PE). We aimed to identify the proteins carried by STBM and their potential pathological roles in early-onset severe PE. METHODS: In this study, we performed a differential proteomic analysis of STBM from early-onset severe PE patients, using iTRAQ isobaric tags and 2D nano LC-MS/MS. STBM were generated by the in vitro explant culture method, and then verified by electron microscopy and western blot analysis. RESULTS: A total of 18 533 unique peptides and 3 317 proteins were identified, 3 292 proteins were quantified. We identified 194 differentially expressed proteins in STBM from early-onset severe PE patients, 122 proteins were up-regulated and 72 proteins were down-regulated. Further bioinformatics analysis revealed that mitochondrion, transmembrane transport and transmembrane transporter activity were the most abundant categories in gene ontology (GO) annotation. Glycolysis/ gluconeogenesis, citrate cycle, fatty acid elongation, steroid hormone biosynthesis and oxidative phosphorylation were the five significantly represented pathways. Four differentially expressed proteins (siglec-6, calnexin, CD63 and S100-A8) related to inflammation, coagulation or immunoregulation were independently verified using western blot. CONCLUSIONS: The identification of key proteins carried by STBM may serve not only as a basis for better understanding and further exploring the etiology and pathogenesis of PE, but also as potential biomarkers and in providing targets for future therapy in PE, especially in early-onset severe PE(sPE).
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Vesículas Extracelulares/química , Pré-Eclâmpsia/genética , Proteoma/genética , Trofoblastos/metabolismo , Adulto , Técnicas de Cultura de Células , Ciclo do Ácido Cítrico/genética , Ácidos Graxos/biossíntese , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Gluconeogênese/genética , Glicólise/genética , Humanos , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Anotação de Sequência Molecular , Pré-Eclâmpsia/metabolismo , Pré-Eclâmpsia/patologia , Gravidez , Proteoma/metabolismo , Índice de Gravidade de Doença , Coloração e Rotulagem , Espectrometria de Massas em Tandem , Fatores de Tempo , Trofoblastos/patologiaRESUMO
Cervical cancer poses a significant health burden for women globally, and the rapid proliferation of cervical cancer cells greatly worsens patient prognosis. Long non-coding RNAs (lncRNAs) play a crucial role in regulating tumor cell proliferation. However, the involvement of lncRNAs in cervical cancer cell proliferation remains unclear. In this study, we investigated the lncRNA SIX1-1, which was found to be upregulated in cervical cancer tissues and cell lines. Functional assays revealed that knockdown of SIX1-1 inhibited cell proliferation in vitro and reduced tumor growth in vivo. Mechanistically, SIX1-1 was predominantly localized in the nucleus and could bind with DNMT1 protein. The expression of SIX1-1 enhanced the interaction of DNMT1 with RASD1 promoter, leading to the methylation of the promoter and decreased mRNA transcription. Then RASD1 downregulation activated the cAMP/PKA/CREB signaling pathway, promoting cell proliferation. Rescue experiments showed that knockdown of RASD1 restored the inhibited cell proliferation caused by decreased expression of SIX1-1, indicating that RASD1 acted as the functional mediator of SIX1-1. In conclusion, SIX1-1 promoted cervical cancer cell proliferation by modulating RASD1 expression. This suggests that targeting the SIX1-1/RASD1 axis could be a potential antitumor strategy for cervical cancer.
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An innovative salamo-like fluorescent chemical sensor H2L, has been prepared that can be utilized to selectively detect Cu2+ and B4O72- ions. Cu2+ ions can bind to oxime state nitrogen and phenol state oxygen atoms in the chemosensor H2L, triggering the LMCT effect leading to fluorescence enhancement. The crystal structure of the copper(II) complex, named as [Cu(L)], has been achieved via X-ray crystallography, and the sensing mechanism has been confirmed by further theoretical calculations with DFT. Besides, the sensor H2L recognizes B4O72- ions causing an ICT effect resulting in bright blue fluorescence. Moreover, the sensor has relatively high selectivity and sensitivity for Cu2+ and B4O72- ions, and the detection limits are 1.02 × 10-7 and 2.06 × 10-7 M, respectively. In addition, the good biocompatibility and excellent water solubility of the sensor H2L make it very advantageous in practical applications, using H2L powder for fingerprint visualization, using H2L to identify the phenomenon of B4O72- ions emitting bright blue fluorescence, making it an ink that can print encrypted messages on A4 paper, in addition to this, based on H2L, the real water sample was tested for Cu2+ ion recognition, and finally the test strip experiment was carried out.
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BACKGROUND: H19 is a paternally imprinted gene that has been shown to be highly expressed in the trophoblast tissue. Results from previous studies have initiated a debate as to whether noncoding RNA H19 acts as a tumor suppressor or as a tumor promotor in trophoblast tissue. In the present study, we developed lentiviral vectors expressing H19-specific small interfering RNA (siRNA) to specifically block the expression of H19 in the human choriocarcinoma cell line JAR. Using this approach, we investigated the impact of the H19 gene on the proliferation, invasion and apoptosis of JAR cells. Moreover, we examined the effect of H19 knockdown on the expression of insulin-like growth factor 2 (IGF2), hairy and enhancer of split homologue-1 (HES-1) and dual-specific phosphatase 5 (DUSP5) genes. RESULTS: H19 knockdown inhibited apoptosis and proliferation of JAR cells, but had no significant impact on cell invasion. In addition, H19 knockdown resulted in significant upregulation of HES-1 and DUSP5 expression, but not IGF2 expression in JAR cells. CONCLUSIONS: The finding that H19 downregulation could simultaneously inhibit proliferation and apoptosis of JAR cells highlights a putative dual function for H19 in choriocarcinoma and may explain the debate on whether H19 acts as a tumor suppressor or a tumor promotor in trophoblast tissue. Furthermore, upregulation of HES-1 and DUSP5 may mediate H19 downregulation-induced suppression of proliferation and apoptosis of JAR cells.
Assuntos
Apoptose/fisiologia , Proliferação de Células , Coriocarcinoma/patologia , Lentivirus/genética , Proteínas Nucleares/genética , Interferência de RNA/fisiologia , Proteínas Supressoras de Tumor/genética , Neoplasias Uterinas/patologia , Apoptose/genética , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Linhagem Celular Tumoral , Coriocarcinoma/metabolismo , Fosfatases de Especificidade Dupla/metabolismo , Feminino , Técnicas de Silenciamento de Genes , Vetores Genéticos/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Fator de Crescimento Insulin-Like II/metabolismo , Proteínas Nucleares/fisiologia , Gravidez , Fatores de Transcrição HES-1 , Proteínas Supressoras de Tumor/fisiologia , Neoplasias Uterinas/metabolismoRESUMO
OBJECTIVE: To investigate the clinical significance of abnormal human telomerase RNA gene component (hTERC) gene amplification tested by fluorescence in situ hybridization in cervical lesions. METHODS: In 373 patients with cytologic abnormalities, high-risk human papilomavirus (HR-HPV) was detected by the hybrid capture II method, and abnormal amplification of the hTERC gene in exfoliated cells was detected by fluorescence in situ hybridization. RESULTS: Cell smear findings suggested atypical squamous cells in 148 patients, low-grade squamous intraepithelial lesion in 62 patients, and high-grade squamous intraepithelial lesion in 107 patients, squamous cell carcinoma in 56 patients, and cervical biopsy-revealed inflammation in 89 patients, cervical intraepithelial neoplasia (CIN) I in 36 patients, CIN II in 43 patients, CIN III in 129 patients, and infiltrating carcinoma in 76 patients. In the inflammation, CIN I, CIN II, CIN III, and infiltrating carcinoma groups, the infection rates of HR-HPV were 29.21%, 52.78%, 74.42%, 92.25%, and 93.42% (P < 0.01), respectively; the positive rates of hTERC gene amplification were 0.00%, 13.89%, 41.86%, 78.29%, and 89.47% (P < 0.01), respectively. With respect to advanced cervical lesions (≥CIN II), cytology (≥ low-grade squamous intraepithelial lesion), HR-HPV testing, and hTERC testing differed insignificantly in the negative predictive value (P > 0.05), but they differed significantly in the sensitivity, specificity, and positive predictive value (P < 0.01). Among the 3 methods, hTERC testing showed the highest specificity and positive predictive value, and HR-HPV testing showed the highest sensitivity. In 41 patients with untreated CIN I and CIN II, the sensitivity of detection of hTERC gene amplification to predict lesion progression was 88.89%, and the specificity was 93.75%. CONCLUSION: Detection of abnormal amplification of the hTERC gene can assist in screening cervical lesions and identifying CIN I/II patients with a high progression risk.
Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/diagnóstico , Amplificação de Genes , Inflamação/diagnóstico , RNA/genética , Telomerase/genética , Displasia do Colo do Útero/diagnóstico , Neoplasias do Colo do Útero/diagnóstico , Adulto , Idoso , Carcinoma de Células Escamosas/genética , Citodiagnóstico , Feminino , Humanos , Hibridização in Situ Fluorescente , Inflamação/genética , Pessoa de Meia-Idade , Gradação de Tumores , Prognóstico , Displasia do Colo do Útero/genética , Neoplasias do Colo do Útero/genética , Adulto JovemRESUMO
Octamer-binding transcription factor 4 (Oct4), an important embryonic transcriptional factor, is highly expressed in several tumors and is considered as a hallmark of cancer stem cells. Knowledge about the expression and regulatory mechanisms of Oct4 can contribute to the treatment of cancers. As for cervical cancer, however, details remain obscure about Oct4 expression and its regulatory mechanism. In this study, we found that the level of Oct4 in human papillomavirus 16 (HPV16)- positive cervical cancer cells (CaSki cells) was higher than that in HPV-negative cervical cancer cells (C-33A cells), whereas both the level of histone deacetylase 1 (HDAC1) and DNA methyltransferase 3A (DNMT3A) were lower in CaSki cells than those in C-33A cells. Treatment with valproic acid, an HDAC inhibitor, could significantly increase the expression of Oct4 in C-33A cells, but only slightly increased Oct4 in CaSki cells. Co-immunoprecipitation assays showed that HDAC1 and DNMT3A existed in a common complex. The co-immunoprecipitated DNMT3A or HDAC1 was dose-dependently decreased with valproic acid treatment. These results indicated that HDAC1/DNMT3A-containing complex is associated with the suppression of Oct4 in cervical cancer cells, and the activity of HDAC1 is required in the repression of Oct4.
Assuntos
DNA (Citosina-5-)-Metiltransferases/metabolismo , Histona Desacetilase 1/metabolismo , Fator 3 de Transcrição de Octâmero/antagonistas & inibidores , Neoplasias do Colo do Útero/metabolismo , DNA Metiltransferase 3A , Feminino , Histona Desacetilase 1/antagonistas & inibidores , Humanos , Fator 3 de Transcrição de Octâmero/biossíntese , Fator 3 de Transcrição de Octâmero/metabolismo , Relação Estrutura-Atividade , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/patologia , Ácido Valproico/farmacologiaRESUMO
The colony-forming ability of cervical cancer is affected by many factors. Oct4, an important transcription factor, is highly expressed in several tumors and promotes the colony-forming ability of cancer cells. Thus, it is considered a potential target for the treatment of cancer. However, we know little about the expression level of Oct4 and its epigenetic regulatory mechanism in cervical cancer cells. In this study, we are the first to observe that human papillomavirus (HPV)-positive cervical cancer cell lines (HeLa, Caski) have a stronger colony-forming ability than HPV-negative cervical cancer cell lines (C-33A). Moreover, the expression level of Oct4 in both HeLa and Caski cells was also higher than that in C-33A cells. We then confirmed that there was a negative correlation between the expression of Oct4 and DNMT3A in these three types of cervical cancer cells, whereas DNA methyltransferase 1 and 3B had no differences among the cell lines. However, after DNA methylation in both key regulatory regions of the Oct4 gene and the genomic levels were analyzed, we found that DNA methyltransferase 3A could neither regulate the expression of Oct4 nor affect the whole level of genomic DNA methylation. These results suggest three points: (1) Oct4 might be treated as a new target for the treatment of cervical cancer, (2) we could not inhibit the expression of Oct4 by DNA demethylation, and (3) HPV virus might initiate cervical carcinogenesis by upregulation of Oct4 expression.
Assuntos
DNA (Citosina-5-)-Metiltransferases/metabolismo , Regulação Neoplásica da Expressão Gênica/genética , Fator 3 de Transcrição de Octâmero/biossíntese , Infecções por Papillomavirus/metabolismo , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/virologia , Western Blotting , Cromatografia Líquida de Alta Pressão , DNA (Citosina-5-)-Metiltransferases/genética , DNA Metiltransferase 3A , Feminino , Imunofluorescência , Células HeLa , Humanos , Fator 3 de Transcrição de Octâmero/genética , Infecções por Papillomavirus/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Neoplasias do Colo do Útero/genéticaRESUMO
HMGB1, composed of the A box, B box, and C tail domains, is a critical proinflammatory cytokine involved in diverse inflammatory diseases. The B box mediates proinflammatory activity, while the A box alone acts as a specific antagonist of HMGB1. The C tail contributes to the spatial structure of A box and regulates HMGB1 DNA binding specificity. It is unknown whether the C tail can enhance the anti-inflammatory effect of A box. In this study, we generated fusion proteins consisting of the A box and C tail, in which the B box was deleted and the A box and C tail were linked either directly or by the flexible linker sequence (Gly4Ser)3. In vitro and in vivo experiments showed that the two fusion proteins had a higher anti-inflammatory activity compared to the A box alone. This suggests that the fused C tail enhances the anti-inflammatory effect of the A box.
Assuntos
Anti-Inflamatórios/metabolismo , Proteína HMGB1/química , Proteína HMGB1/metabolismo , Proteínas Recombinantes de Fusão/metabolismo , Animais , Western Blotting , Linhagem Celular , Eletroforese em Gel de Poliacrilamida , Endotoxinas/toxicidade , Humanos , Interleucina-6/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/química , Fator de Necrose Tumoral alfa/metabolismoRESUMO
INTRODUCTION: The homeobox gene Six1 is overexpressed in multiple human tumors, playing a role in promoting tumorigenesis and metastasis. The present study was aimed to investigate the clinical implications of Six1 expression in cervical cancer. METHODS: Six1 messenger RNA (mRNA) and protein expression was detected by reverse transcription (RT) polymerase chain reaction and Western blotting, respectively, in human cervical cancer cell lines CaSki, HeLa, C33A and 20 normal cervical specimens, 21 specimens of cervical intraepithelial neoplasias (CINs), and 54 specimens of cervical cancer tissue, and the clinical implications of Six1 gene expression was analyzed. RESULTS: There was Six1 mRNA and protein overexpression in cervical cancer cell lines CaSki, HeLa, and C33A. The Six1 expression level was higher in CaSki and HeLa cells than in C33A cells (P < 0.05). Six1 mRNA and protein expression increased from normal cervical epithelial tissues, to CINs, and then to cervical cancer tissue (normal cervical epithelial tissue vs CIN, P < 0.05; normal cervical epithelial tissue vs cervical cancer, and CIN vs cervical cancer, P < 0.01). The status of Six1 overexpression was correlated to clinical staging and lymph node metastasis of cervical cancer (P < 0.01) but not to pathological grading, tumor size, and age of the patient (P > 0.05). CONCLUSION: Six1 was overexpressed in cervical cancer cell lines and in cervical cancer tissues. Alteration of Six1 expression might contribute to the occurrence and development of cervical cancer.
Assuntos
Colo do Útero/metabolismo , Epitélio/metabolismo , Proteínas de Homeodomínio/genética , Displasia do Colo do Útero/genética , Neoplasias do Colo do Útero/genética , Adulto , Idoso , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Colo do Útero/patologia , Epitélio/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Genes Homeobox , Células HeLa , Proteínas de Homeodomínio/metabolismo , Humanos , Pessoa de Meia-Idade , Neoplasias do Colo do Útero/metabolismo , Neoplasias do Colo do Útero/patologia , Displasia do Colo do Útero/metabolismo , Displasia do Colo do Útero/patologiaRESUMO
OBJECTIVES: It was the aim of this study to investigate the feasibility of polymerase chain reaction (PCR)/ligase detection reaction (LDR)/capillary electrophoresis for the detection of paternally inherited fetal CD17 (A-->T) mutation of beta-thalassemia in maternal plasma. METHODS: The target DNA in maternal plasma was amplified by PCR and the mutant signal was detected by LDR. Unique LDR products were produced and separated by capillary electrophoresis. PCR/LDR/capillary electrophoresis was applied to detect CD17 (A-->T) mutation in an experimental model at different sensitivity levels and from 3 maternal plasma samples. RESULTS: The sensitivity of PCR/LDR/capillary electrophoresis for detecting low-abundance CD17 (A-->T) mutation was 1:5,000 at least. The technique was applied in maternal plasma DNA for detecting paternally inherited fetal CD17 (A-->T) mutation, and the results were concordant with that of PCR/reverse dot blot of amniotic fluid cell DNA. CONCLUSIONS: PCR/LDR/capillary electrophoresis has a very high sensitivity to distinguish low-abundance single nucleotide differences and probably detects paternally inherited fetal point mutations in maternal plasma.
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Análise Mutacional de DNA , DNA/sangue , Diagnóstico Pré-Natal/métodos , Talassemia beta/diagnóstico , Eletroforese Capilar , Feminino , Feto , Testes Genéticos/métodos , Genótipo , Humanos , Troca Materno-Fetal , Mutação , Reação em Cadeia da Polimerase , Gravidez , Talassemia beta/genéticaRESUMO
B cell activating factor belonging to the TNF family (BAFF) is a novel member of the tumor necrosis factor (TNF) ligand family and plays an important role in B lymphocyte maturation and survival. Overexpression of BAFF is closely involved in the pathogenesis and progression of many kinds of autoimmune disorders; therefore, BAFF has been considered as an ideal therapeutic target for these conditions. In this study, we generated several candidate immune inhibitors of human BAFF by conjugating foreign immunodominant T-helper cell (Th) epitopes to the N- or C-terminus of five BAFF mutants. The recombined proteins were successfully expressed in Escherichia coli (E. coli) and purified by Ni-NTA chromatography. BALB/c mice immunized with the recombinant proteins produced high levels of anti-BAFF antibodies, and their sera inhibited the lymphocyte proliferation-inducing activity of recombinant soluble BAFF and natural soluble BAFF. Moreover, antibodies cross-reactive with BAFF were detected in sera from hu-SCID mice immunized with the recombinant proteins. These results indicated that the recombinant BAFF mutants modified with Th epitopes could induce neutralizing antibodies against BAFF in vivo. This study may provide a valuable strategy for treating BAFF-associated autoimmune diseases.
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Anticorpos/farmacologia , Fator Ativador de Células B/antagonistas & inibidores , Fator Ativador de Células B/imunologia , Proteínas Mutantes/imunologia , Animais , Anticorpos/sangue , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Imunização , Imunoglobulina G/sangue , Transfusão de Linfócitos , Linfócitos/citologia , Linfócitos/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos SCID , Testes de Neutralização , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/isolamento & purificação , SolubilidadeRESUMO
Dichloroacetate (DCA), a traditional mitochondria-targeting agent, has shown promising prospect as a sensitizer in fighting against malignancies including cervical cancer. But it is unclear about the effect of DCA alone on cervical tumor. Moreover, previous reports have demonstrated that the increased cyclooxygenase-2 (COX2) expression is associated with chemoresistance and poor prognosis of cervical cancer. However, it is still unknown whether COX2 can affect the sensitivity of DCA in cervical cancer cells. In this study, we found that cervical cancer cells were insensitive to DCA. Furthermore, we for the first time revealed that DCA could upregulate COX2 which impeded the chemosensitivity of DCA in cervical cancer cells. Mechanistic study showed that DCA reduced the level of RNA binding protein quaking (QKI), leading to the decay suppression of COX2 mRNA and the subsequent elevation of COX2 protein. Inhibition of COX2 using celecoxib could sensitize DCA in repressing the growth of cervical cancer cells both in vitro and in vivo. These results indicate that COX2 is a novel resistance factor of DCA, and combination of celecoxib with DCA may be beneficial to the treatment of cervical cancer.
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Pirarubicin (THP) is a newer generation anthracycline anticancer drug. In the clinic, THP and THP-based combination therapies have been demonstrated to be effective against various tumors without severe side effects. However, previous clinical studies have shown that most patients with cervical cancer are not sensitive to THP treatment, and the associated mechanisms are not clear. Consistent with the clinical study, we confirmed that cervical cancer cells were resistant to THP in vitro and in vivo. Our data demonstrated that THP induced a protective macroautophagy/autophagy response in cervical cancer cells, and suppression of this autophagy dramatically enhanced the cytotoxicity of THP. By scanning the mRNA level change of autophagy-related genes, we found that the upregulation of ATG4B (autophagy-related 4B cysteine peptidase) plays an important role in THP-induced autophagy. Moreover, THP increased the mRNA level of ATG4B in cervical cancer cells by promoting mRNA stability without influencing its transcription. Furthermore, THP triggered a downregulation of MIR34C-5p, which was associated with the upregulation of ATG4B and autophagy induction. Overexpression of MIR34C-5p significantly decreased the level of ATG4B and attenuated autophagy, accompanied by enhanced cell death and apoptosis in THP-treated cervical cancer cells. These results for the first time reveal the presence of a MIR34C-5p-ATG4B-autophagy signaling axis in THP-treated cervical cancer cells in vitro and in vivo, and the axis, at least partially, accounts for the THP nonsensitivity in cervical cancer patients. This study may provide a new insight for improving the chemotherapeutic effect of THP, which may be beneficial to the further clinical application of THP in cervical cancer treatment.
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Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Proteínas Relacionadas à Autofagia/metabolismo , Autofagia/efeitos dos fármacos , Cisteína Endopeptidases/metabolismo , Doxorrubicina/análogos & derivados , MicroRNAs/genética , Autofagia/genética , Linhagem Celular Tumoral , Doxorrubicina/farmacologia , Feminino , Humanos , Transdução de Sinais/efeitos dos fármacos , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/genéticaRESUMO
Hypertension is a major risk factor for cardiovascular and cerebrovascular disease. Prenatal exposure to lipopolysaccharide (LPS) leads to hypertension in a rat offspring. However, the mechanism is still unclear. This study unraveled epigenetic mechanism for this and explored the protective effects of ascorbic acid against hypertension on prenatal inflammation-induced offspring. Prenatal LPS exposure resulted in an increase of intrarenal oxidative stress and enhanced angiotensin-converting enzyme 1 (ACE1) gene expression at the mRNA and protein levels in 6- and 12-week-old offspring, correlating with the augmentation of histone H3 acetylation (H3AC) on the ACE1 promoter. However, the prenatal ascorbic acid treatment decreased the LPS-induced expression of ACE1, protected against intrarenal oxidative stress, and reversed the altered histone modification on the ACE1 promoter, showing the protective effect in offspring of prenatal LPS stimulation. Our study demonstrates that ascorbic acid is able to prevent hypertension in offspring from prenatal inflammation exposure. Thus, ascorbic acid can be a new approach towards the prevention of fetal programming hypertension.
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Ácido Ascórbico/farmacologia , Regulação da Expressão Gênica no Desenvolvimento , Histonas/metabolismo , Hipertensão/prevenção & controle , Peptidil Dipeptidase A/metabolismo , Acetilação , Animais , Antioxidantes/metabolismo , Pressão Sanguínea , Peso Corporal , Ilhas de CpG , Epigênese Genética , Feminino , Hipertensão/metabolismo , Inflamação/metabolismo , Rim/patologia , Lipopolissacarídeos/metabolismo , Estresse Oxidativo , Peptidil Dipeptidase A/genética , Gravidez , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Regiões Promotoras Genéticas , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Fatores de RiscoRESUMO
Both dichloroacetate (DCA) and metformin (Met) have shown promising antitumor efficacy by regulating cancer cell metabolism. However, the DCA-mediated protective autophagy and Met-induced lactate accumulation limit their tumor-killing potential respectively. So overcoming the corresponding shortages will improve their therapeutic effects. In the present study, we found that DCA and Met synergistically inhibited the growth and enhanced the apoptosis of ovarian cancer cells. Interestingly, we for the first time revealed that Met sensitized DCA via dramatically attenuating DCA-induced Mcl-1 protein and protective autophagy, while DCA sensitized Met through markedly alleviating Met-induced excessive lactate accumulation and glucose consumption. The in vivo experiments in nude mice also showed that DCA and Met synergistically suppressed the growth of xenograft ovarian tumors. These results may pave a way for developing novel strategies for the treatment of ovarian cancer based on the combined use of DCA and Met.
Assuntos
Antineoplásicos/uso terapêutico , Ácido Dicloroacético/uso terapêutico , Inibidores do Crescimento/uso terapêutico , Metformina/uso terapêutico , Neoplasias Ovarianas/tratamento farmacológico , Animais , Apoptose , Autofagia , Linhagem Celular Tumoral , Sinergismo Farmacológico , Quimioterapia Combinada , Feminino , Humanos , Ácido Láctico/metabolismo , Camundongos , Camundongos Nus , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: To determine the diagnostic accuracy, validity, current limitations of, and possible solutions to, fetal RhD genotyping from maternal blood based on existing studies written in English. METHODS: A literature search was conducted that described fetal RhD determination from maternal blood. The number of samples tested, fetal RhD genotype, the source of cell-free fetal DNA, gestational age and fetal Rh type were examined in each study to calculate the accuracy, sensitivity and specificity of fetal RhD genotyping. RESULTS: Forty-one publications, which included 11,129 samples with non-invasive Rh genotyping of cell-free fetal DNA from maternal blood, were selected. After the exclusion of 352 inconclusive samples, the overall diagnostic accuracy was 98.5% (10,611/10,777), and sensitivity and specificity were 99% and 98%, respectively. First trimester diagnosis showed an accuracy of 99%, higher than second and third trimester diagnosis. Thirty studies reported a 100% diagnostic accuracy of fetal RhD genotyping. CONCLUSION: Non-invasive fetal RhD genotyping from maternal blood has high accuracy, sensitivity and specificity. METHODS reducing false results have been explored and applied in research. These achievements indicate that this technique will be widely used in routine clinical care.
Assuntos
DNA/sangue , Técnicas de Genotipagem/métodos , Gravidez/sangue , Diagnóstico Pré-Natal/métodos , Sistema do Grupo Sanguíneo Rh-Hr/genética , Sistema Livre de Células , DNA/análise , Eritroblastose Fetal/diagnóstico , Eritroblastose Fetal/genética , Feminino , Feto/metabolismo , Humanos , Recém-Nascido , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
BACKGROUND: Preeclampsia (PE) and eclampsia remain leading causes of maternal and fetal mortality worldwide. The kidney is considered the first and most severely affected organ in women with PE/eclampsia. In this study, we analyzed new morphologic features of kidney biopsies and clinical findings in patients with PE or eclampsia at our hospital. METHODS: Eight patients with PE/eclampsia underwent renal biopsies during the antepartum (3/8) or postpartum (5/8) period. Maternal clinical findings, major serological indices, neonatal outcomes, and renal histopathologic and immunofluorescent characteristics were reviewed for each case. RESULTS: Most patients had abnormal serum cholesterol (8/8), triglyceride (6/8), albumin (7/8), and uric acid (5/8). The ratio of blood urea nitrogen (BUN) to serum creatinine (SCr) was elevated in all patients. Five of eight newborns survived. Various degrees of morphologic change were present in the renal glomeruli, and were associated with proteinuria. All patients had deposition of complement factor 4 (C4) in the renal glomeruli and seven had deposition of immunoglobulin M (IgM). CONCLUSION: Endotheliosis, vacuolation of podocytes, proliferation of mesangial cells, and protein casts in the tubule lumens were found in the kidneys of women with PE/eclampsia. Immune depositions of C4 and IgM are major contributors to renal lesions in preeclamptic patients, whose neonates can generally survive. Eclampsia can occur without increased blood pressure.