RESUMO
Emerging evidence has suggested that N6 -methyladenosine (m6 A) regulates the pathology of Parkinson's disease (PD). Nevertheless, the function of demethylase fat mass and obesity (FTO) associated pathogenesis is still not fully elucidated. Here, this research findings revealed that m6 A-modification was decreased in PD models, meanwhile, the FTO level upregulated in the PD models. Functionally, in N-methyl-4-phenylpyridinium (MPP+) treated SH-SY5Y cells, the ferroptosis significantly upregulated and FTO silencing mitigated the ferroptosis phenotype. Moreover, in silico assays indicated that nuclear factor erythroid 2-related factor-2 (NRF2) acted as the target of FTO, and FTO demethylated the m6 A modification from NRF2 mRNA. Furthermore, FTO impaired the NRF2 mRNA stability via m6 A-dependent pathway. Thus, our findings illustrated an important role of FTO on PD through m6 A-NRF2-ferroptosis manner. Taken together, the study revealed the potential function of FTO on PD nervous system diseases.
Assuntos
Adenina/análogos & derivados , Ferroptose , Neuroblastoma , Doença de Parkinson , Humanos , Doença de Parkinson/genética , Fator 2 Relacionado a NF-E2/genética , Obesidade/genética , 1-Metil-4-fenilpiridínio , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genéticaRESUMO
17ß-Estradiol (E2) is an important endogenous estrogen, which disturbs the endocrine system and poses a threat to human health because of its accumulation in the human body. Herein, a biofuel cell (BFC)-based self-powered electrochemical aptasensor was developed for E2 detection. Porous carbon nanocage/gold nanoparticle composite modified indium tin oxide (CNC/AuNP/ITO) and glucose oxidase modified CNC/AuNP/ITO were used as the biocathode and bioanode of BFCs, respectively. [Fe(CN)6]3- was selected as an electroactive probe, which was entrapped in the pores of positively charged magnetic Fe3O4 nanoparticles (PMNPs) and then capped with a negatively charged E2 aptamer to form a DNA bioconjugate. The presence of the target E2 triggered the entrapped [Fe(CN)6]3- probe release due to the removal of the aptamer via specific recognition, which resulted in the transfer of electrons produced by glucose oxidation at the bioanode to the biocathode and produced a high open-circuit voltage (EOCV). Consequently, a "signal-on" homogeneous self-powered aptasensor for E2 assay was realized. Promisingly, the BFC-based self-powered aptasensor has particularly high sensitivity for E2 detection in the concentration range of 0.5 pg mL-1 to 15 ng mL-1 with a detection limit of 0.16 pg mL-1 (S/N = 3). Therefore, the proposed BFC-based self-powered electrochemical aptasensor has great promise to be applied as a successful prototype of a portable and on-site bioassay in the field of environment monitoring and food safety.
Assuntos
Aptâmeros de Nucleotídeos , Fontes de Energia Bioelétrica , Carbono , Técnicas Eletroquímicas , Estradiol , Ouro , Nanopartículas Metálicas , Estradiol/química , Estradiol/análise , Aptâmeros de Nucleotídeos/química , Ouro/química , Nanopartículas Metálicas/química , Carbono/química , Técnicas Eletroquímicas/métodos , Técnicas Eletroquímicas/instrumentação , Técnicas Biossensoriais/métodos , Limite de Detecção , Humanos , DNA/química , Glucose Oxidase/química , Compostos de Estanho/químicaRESUMO
T4 polynucleotide kinase helps with DNA recombination and repair. In this study, an electrochemical biosensor was developed for a T4 polynucleotide kinase activity assay and inhibitor screening based on phosphate pillar[5]arene and multi-walled carbon nanotube nanocomposites. The water-soluble pillar[5]arene was employed as the host to complex thionine guest molecules. The substrate DNA with a 5'-hydroxyl group initially self-assembled on the gold electrode surface through chemical adsorption of the thiol group, which was phosphorylated in the presence of T4 polynucleotide kinase. Titanium dioxide nanoparticles served as a bridge to link phosphorylated DNA and phosphate pillar[5]arene and multi-walled carbon nanotube composite due to strong phosphate-Ti4+-phosphate chemistry. Through supramolecular host-guest recognition, thionine molecules were able to penetrate the pillar[5]arene cavity, resulting in an enhanced electrochemical response signal. The electrochemical signal is proportional to the T4 polynucleotide kinase concentration in the range of 10-5 to 15 U mL-1 with a detection limit of 5 × 10-6 U mL-1. It was also effective in measuring HeLa cell lysate-related T4 polynucleotide kinase activity and inhibitor screening. The proposed method offers a unique sensing platform for kinase activity measurement, holding great potential in nucleotide kinase-target drug development, clinical diagnostics, and inhibitor screening.
Assuntos
Técnicas Biossensoriais , Nanotubos de Carbono , Fenotiazinas , Humanos , Polinucleotídeo 5'-Hidroxiquinase , Nanotubos de Carbono/química , Fosfatos , Células HeLa , DNA/química , Técnicas Biossensoriais/métodosRESUMO
A novel biofuel cell (BFC)-based self-powered electrochemical immunosensing platform was developed by integrating the target-induced biofuel release and biogate immunoassay for ultrasensitive 17ß-estradiol (E2) detection. The carbon nanocages/gold nanoparticle composite was employed in the BFCs device as the electrode material, through which bilirubin oxidase and glucose oxidase were wired to form the biocathode and bioanode, respectively. Positively charged mesoporous silica nanoparticles (PMSN) were encapsulated with glucose molecules as biofuel and subsequently coated by the negatively charged AuNPs-labelled anti-E2 antibody (AuNPs-Ab) serving as a biogate. The biogate could be opened efficiently and the trapped glucose released once the target E2 was recognized and captured by AuNPs-Ab due to the decreased adhesion between the antigen-antibody complex and PMSN. Then, glucose oxidase oxidized the glucose to produce a large number of electrons, resulting in significantly increased open-circuit voltage (EOCV). Promisingly, the proposed BFC-based self-powered immunosensor demonstrated exceptional sensitivity for the detection of E2 in the concentration range from 1.0 pg mL-1 to 10.0 ng mL -1, with a detection limit of 0.32 pg mL-1 (S/N = 3). Furthermore, the prepared BFC-based self-powered homogeneous immunosensor showed significant potential for implementation as a viable prototype for a mobile and an on-site bioassay system in food and environmental safety applications.
Assuntos
Fontes de Energia Bioelétrica , Técnicas Biossensoriais , Estradiol , Glucose Oxidase , Ouro , Limite de Detecção , Nanopartículas Metálicas , Imunoensaio/métodos , Estradiol/química , Estradiol/análise , Ouro/química , Glucose Oxidase/química , Técnicas Biossensoriais/métodos , Nanopartículas Metálicas/química , Técnicas Eletroquímicas/métodos , Técnicas Eletroquímicas/instrumentação , Humanos , Eletrodos , Glucose/análise , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/química , Anticorpos Imobilizados/imunologia , Dióxido de Silício/química , Enzimas Imobilizadas/químicaRESUMO
All-polymer photodetectors possess unique mechanical flexibility and are ideally suitable for the application in next-generation flexible, wearable short-wavelength infrared (SWIR, 1000-2700 nm) photodetectors. However, all-polymer photodetectors commonly suffer from low sensitivity, high noise, and low photoresponse speed in the SWIR region, which significantly diminish their application potential in wearable electronics. Herein, two polymer acceptors with absorption beyond 1000 nm, namely P4TOC-DCBT and P4TOC-DCBSe, were designed and synthesized. The two polymers possess rigid structure and good conformational stability, which is beneficial for reducing energetic disorder and suppressing dark current. Owing to the efficient charge generation and ultralow noise current, the P4TOC-DCBT-based all-polymer photodetector achieved a specific detectivity () of over 1012 Jones from 650 (visible) to 1070 nm (SWIR) under zero bias, with a response time of 1.36 µs. These are the best results for reported all-polymer SWIR photodetectors in photovoltaic mode. More significantly, the all-polymer blend films exhibit good mechanical durability, and hence the P4TOC-DCBT-based flexible all-polymer photodetectors show a small performance attenuation (<4%) after 2000 cycles of bending to a 3 mm radius. The all-polymer flexible SWIR organic photodetectors are successfully applied in pulse signal detection, optical communication and image capture.
RESUMO
PURPOSE: Here, we aim to identify a CEACAM5-targeted nanobody and demonstrate its application in positron emission tomography (PET) imaging and near-infrared (NIR) fluorescence imaging in colorectal cancer (CRC). METHODS: Immunohistochemistry was applied to verify CEACAM5 expression in CRC and metastatic lymph nodes (mLNs). CEACAM5-targeted nanobodies were obtained by immunization of human CEACAM5 protein in a dromedary, followed by several rounds of phage screenings. Immunofluorescence staining and flow cytometry was carried out to determine the binding affinity of the nanobodies. The nanobodies were radiolabeled by coupling 18F-SFB for PET imaging of CRC subcutaneous xenografts and lymph node metastasis (LNM). IRDye800CW (IR800) were conjugated to form NIR probes for NIR imaging in CRC subcutaneous models. RESULTS: CEACAM5 was overexpressed in either human CRC tissues or mLNs. A CEACAM5 targeted nanobody, Nb41 was successfully generated, with excellent in vitro binding properties. Incorporation of albumin binding domain (ABD) did not affect the affinity of Nb41. In vivo imaging showed that both 18F-FB-Nb41 and 18F-FB-Nb41-ABD showed obvious accumulation in the tumor. Due to the longer retention in the blood, 18F-FB-Nb41-ABD enrichment in tumors was significantly delayed but higher compared to 18F-FB-Nb41. Both 18F-FB-Nb41 and 18F-FB-Nb41-ABD showed prominent LNM enrichment. Similarly, the IR800-conjugated nanobodies Nb41-IR800 and Nb41-ABD-IR800 exhibited superior imaging effects in subcutaneous models, while Nb41-ABD-IR800 exhibited higher fluorescence intensity in the tumor accompanied with a remarkedly delay compared to Nb41-IR800. CONCLUSION: Collectively, we presented the identification and in vivo validation of a CEACAM5-targeted nanobody and a fused nanobody with an ABD, which enabled to the non-invasive visualization of malignancy of CRC using PET imaging and NIR imaging in subcutaneous models as well as LNM models.
Assuntos
Neoplasias Colorretais , Anticorpos de Domínio Único , Humanos , Linhagem Celular Tumoral , Anticorpos de Domínio Único/metabolismo , Tomografia por Emissão de Pósitrons , Neoplasias Colorretais/diagnóstico por imagem , Imagem Óptica , Antígeno Carcinoembrionário , Proteínas Ligadas por GPIRESUMO
T4 polynucleotide kinase (T4 PNK) helps with DNA recombination and repair. In this work, a phosphate pillar[5]arene@palladium nanoparticles@reduced graphene oxide nanocomposite (PP5@PdNPs@rGO)-based electrochemical biosensor was created to identify T4 PNK activities. The PP5 used to complex toluidine blue (TB) guest molecules is water-soluble. With T4 PNK and ATP, the substrate DNA, which included a 5'-hydroxyl group, initially self-assembled over the gold electrode surface by chemical adsorption of the thiol units. Strong phosphate-Zr4+-phosphate chemistry allowed Zr4+ to act as a bridge between phosphorylated DNA and PP5@PdNPs@rGO. Through a supramolecular host-guest recognition connection, TB molecules were able to penetrate the PP5 cavity, where they produced a stronger electrochemical response. With a 5 × 10-7 U mL-1 detection limit, the electrochemical signal is linear in the 10-6 to 1 U mL-1 T4 PNK concentration range. It was also effective in measuring HeLa cell lysate-related PNK activities and screening PNK inhibitors. Nucleotide kinase-target drug development, clinical diagnostics, and screening for inhibitors all stand to benefit greatly from the suggested technology, which offers a unique sensing mechanism for kinase activity measurement.
Assuntos
Nanopartículas Metálicas , Nanocompostos , Humanos , Paládio , Fosfatos , Células HeLa , Polinucleotídeo 5'-Hidroxiquinase , Cloreto de TolônioRESUMO
BACKGROUND: This study aims to investigate the value of the AngioJet thrombectomy system with adjunct of catheter-directed thrombolysis (CDT) in treating lower extremity deep venous thrombosis (LEDVT). METHODS: 48 patients who were clinically confirmed LEDVT and treated by percutaneous mechanical thrombectomy (PMT) combined with CDT, were included in this retrospective study (AJ-CDT, n = 33; Suction-CDT, n = 15). Baseline characteristics, clinical outcomes and surveillance data were reviewed and analyzed. RESULTS: The overall clot reduction rate of AJ-CDT group was significantly higher than that of Suction-CDT group (77.86% vs 64.47%, P = .027). The CDT therapeutic time (5.75 ± 3.04 vs 7.67 ± 2.82 days, P = .045) and urokinase dosage (3.63 ± 2.16 vs 5.76 ± 2.12 million IU, P = .003) were lower in AJ-CDT group, respectively. There was statistical significance in the transient hemoglobinuria between 2 groups (72.73% vs 6.67%, P < .001). At postoperative 48 hours, the serum creatinine (Scr) value was higher in AJ-CDT group compared to Suction-CDT group statistically (78.56 ± 32.16 vs 60.21 ± 15.72 µmol/l, P = .049). However, the incidence of acute kidney injury (AKI) and uric acid (UA) concentration at postoperative 48 hours between these 2 groups were no statistical difference. There was no statistical significance in the Villalta score and post-thrombosis syndrome (PTS) incidence during postoperative follow-up. CONCLUSIONS: AngioJet thrombectomy system is more effective for the treatment of LEDVT by providing a higher clot reduction rate with shorter thrombolytic time and lower thrombolytic drug dosage. However, the device-related potential risk of renal function injury should be taken appropriate precautions.
Assuntos
Terapia Trombolítica , Trombose Venosa , Humanos , Terapia Trombolítica/efeitos adversos , Estudos Retrospectivos , Sucção , Resultado do Tratamento , Trombectomia/efeitos adversos , Trombose Venosa/tratamento farmacológico , Trombose Venosa/cirurgia , Fibrinolíticos/uso terapêutico , Fibrinolíticos/efeitos adversos , Catéteres , Extremidade InferiorRESUMO
BACKGROUND: We aimed to compare the clinical outcomes of pre-emptive angioplasty versus post-thrombotic percutaneous endovascular restoration of dysfunctional arteriovenous fistula (AVF). METHODS: This retrospective study reviewed the data from 80 patients who underwent 114 endovascular interventions for a malfunctioning AVF from July 2016 to August 2019. Stenotic AVFs were treated with pre-emptive angioplasty. Thrombosed AVFs were treated with percutaneous pharmacomechanical fibrinolysis with urokinase used only during the operation or continuously infused. The differences in patency rates were evaluated using the Kaplan-Meier method. In addition, univariate and multivariate regression Cox models were used to determine influential factors on the postintervention primary patency. RESULTS: Post-thrombotic interventions and pre-emptive angioplasty yielded statistically similar rates in clinical success (100 vs. 100%), anatomic success (94 vs. 89%; P = 0.52), complication (4 vs. 11%; P = 0.29), as well as postintervention primary, assisted primary and secondary patency (P = 0.80; 0.57; 0.57). The use of pre-emptive angioplasty was associated with reduced total cost (¥25,108 vs. ¥30,833, P < 0.001). The patients who used urokinase only during the operation prolonged both the primary and assisted primary patency (P = 0.02; 0.002), while those with continuous infusion of urokinase had worst patency rates and high costs (¥39,275 vs. ¥25,108 vs. ¥27,140, P < 0.001). Compared with the other locations, dysfunction in the anastomotic or juxta-anastomotic segment (HR = 0.41, P = 0.001) was associated with prolonged postintervention primary patency. CONCLUSIONS: No clinical outcome differences were found between the post-thrombotic percutaneous endovascular interventions and pre-emptive angioplasty. However, pre-emptive angioplasty decreased access expenditure.
Assuntos
Angioplastia com Balão , Fístula Arteriovenosa , Derivação Arteriovenosa Cirúrgica , Trombose , Angioplastia/efeitos adversos , Angioplastia/métodos , Angioplastia com Balão/efeitos adversos , Angioplastia com Balão/métodos , Fístula Arteriovenosa/etiologia , Derivação Arteriovenosa Cirúrgica/efeitos adversos , Derivação Arteriovenosa Cirúrgica/métodos , Oclusão de Enxerto Vascular/diagnóstico por imagem , Oclusão de Enxerto Vascular/etiologia , Oclusão de Enxerto Vascular/terapia , Humanos , Diálise Renal/efeitos adversos , Estudos Retrospectivos , Trombose/etiologia , Resultado do Tratamento , Ativador de Plasminogênio Tipo Uroquinase/efeitos adversos , Grau de Desobstrução VascularRESUMO
PURPOSE: To identify the spectrum of RB1 gene mutations in 114 Chinese patients with retinoblastoma. METHODS: Genomic DNA was extracted from the peripheral blood of 114 Rb patients. Polymerase chain reactions (PCRs) followed by direct Sanger sequencing were used to screen for mutations in the RB1 gene, which contains 26 exons with flanking intronic sequences, except exon 15. Clinical data, including gender, age at diagnosis, laterality of ocular lesions, and associated symptoms, were recorded and compared. RESULTS: We identified five novel mutations in the RB1 gene. Twenty-five other mutations found in this study have been previously reported. A higher rate of RB1 mutations, with 47.3% of mutations among bilaterally affected patients vs. 6.8% within unilaterally affected patients, was also observed (p < 0.0001). Bilaterally affected patients were diagnosed earlier when compared to unilaterally affected patients (11 ± 7 months versus 20 ± 14 months, p = 0.0002). Furthermore, nonsense mutations were abundant (n = 14), followed by frameshift mutations (n = 8), splicing site mutations (n = 5), while missense mutations were few (n = 3). CONCLUSIONS: We found five novel mutations in RB1 genes, which expands the mutational spectrum of the gene. Children with bilateral Rb exhibited higher mutation rates and were diagnosed earlier than those with unilateral Rb. These findings will inform clinical diagnosis and genetic therapeutic targeting in Rb patients.
Assuntos
Neoplasias da Retina , Retinoblastoma , Humanos , Retinoblastoma/diagnóstico , Códon sem Sentido , Análise Mutacional de DNA , Estudos de Associação Genética , Mutação , Neoplasias da Retina/diagnóstico , China/epidemiologia , Ubiquitina-Proteína Ligases/genética , Proteínas de Ligação a Retinoblastoma/genéticaRESUMO
Sulfiredoxin 1 (SRXN1) is a pivotal regulator of the antioxidant response in eukaryotic cells. However, the role of SRXN1 in hepatocellular carcinoma (HCC) is far from clear. The present study aims to elucidate whether SRXN1 participates in tumorigenesis and metastasis of HCC and to determine the molecular mechanisms. We found that SRXN1 expression was up-regulated in HCC tissue samples and correlated with poor prognosis in HCC patients. We also observed that SRXN1 knockdown by transient siRNA transfection inhibited HCC cell proliferation, migration and invasion. Overexpression of SRXN1 increased HCC cell migration and invasion. B-cell translocation gene 2 (BTG2) was identified as a downstream target of SRXN1. Mechanistic studies revealed that SRXN1-depleted reactive oxygen species (ROS) modulated migration and invasion of HCC cells. In addition, the ROS/p65/BTG2 signalling hub was found to regulate the epithelial-mesenchymal transition (EMT), which mediates the pro-metastasis role of SRXN1 in HCC cells. In vivo experiments showed SRXN1 promotes HCC tumour growth and metastasis in mouse subcutaneous xenograft and metastasis models. Collectively, our results revealed a novel pro-tumorigenic and pro-metastatic function of SRXN1 in HCC. These findings demonstrate a rationale to exploit SRXN1 as a therapeutic target effectively preventing metastasis of HCC.
Assuntos
Carcinoma Hepatocelular/patologia , Transição Epitelial-Mesenquimal/fisiologia , Proteínas Imediatamente Precoces/fisiologia , Neoplasias Hepáticas/patologia , Metástase Neoplásica/fisiopatologia , Proteínas de Neoplasias/fisiologia , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Proteínas Supressoras de Tumor/fisiologia , Animais , Carcinoma Hepatocelular/enzimologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/secundário , Linhagem Celular Tumoral , Movimento Celular , Transformação Celular Neoplásica/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/genética , Neoplasias Pulmonares/secundário , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Invasividade Neoplásica , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/antagonistas & inibidores , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/genética , Interferência de RNA , RNA Mensageiro/genética , RNA Neoplásico/genética , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Fatores de Transcrição/metabolismo , Ensaio Tumoral de Célula-TroncoRESUMO
Cysteinyl leukotrienes (CysLTs) are a group of eicosanoids that regulate the pathogenesis of various human diseases, mainly by signaling through the cysteinyl leukotriene receptor 1 (CysLTR1). The aim of this study was to generate and examine the phenotype of CysLTR1 L118F mutant mice. CysLTR1 L118F mutant mice were generated by the simultaneous microinjection of single guide RNA, Cas9 messenger RNA, and donor plasmid into fertilized mouse eggs. The morphological and behavioral characteristics of the resultant CysLTR1 L118F mutant mice were analyzed using an animal phenotype analysis platform, which included the assessment of body length, tail length, grip strength, and locomotor activity. Immunoprecipitation coupled with mass spectrometry was performed to identify CysLTR1-interacting proteins, and the intracellular calcium levels were determined using fluorometric imaging plate reader assays. The body length and tail length of CysLTR1 L118F mutant mice were significantly increased compared with wild-type mice. In addition, the grip strength and locomotor activity were remarkably elevated in L118F mutant mice compared with wild-type mice. Only three proteins were found to interact with both wild-type and CysLTR1 L118F proteins, whereas 4 and 13 additional proteins interacted exclusively with wild-type and mutant CysLTR1, respectively. Lastly, the responsiveness of cardiac muscle cells to CysLTs were significantly impaired by the L118F substitution in CysLTR1 proteins. The CysLTR1 L118F point mutation induced significant changes in the mouse morphology and behavior, which might be mediated by alterations of its protein interaction profile.
Assuntos
Comportamento Animal , Cálcio/metabolismo , Proteínas Mutantes/metabolismo , Mutação , Miocárdio/metabolismo , Domínios e Motivos de Interação entre Proteínas , Receptores de Leucotrienos/metabolismo , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas Mutantes/genética , Fenótipo , Receptores de Leucotrienos/genéticaRESUMO
Vascular malformations present diagnostic and treatment challenges. In particular, malformations of vessels to the viscera are often diagnosed late or incorrectly due to the insidious onset and deep location of the disease. Therefore, a better knowledge of the genetic mutations underlying such diseases is needed. Here, we evaluated a four-generation family carrying vascular malformations of major vessels that affect multiple organs, which we named "multiorgan venous and lymphatic defect" (MOVLD) syndrome. Genetic analyses identified an association between a mutation in DEAD-box helicase 24 (DDX24), a gene for which the function is largely unknown, and MOVLD. Next, we screened 161 patients with sporadic vascular malformations of similar phenotype to our MOVLD family and found the same mutation or one of the two additional DDX24 mutations in 26 cases. Structural modeling revealed that two of the mutations are located within the adenosine triphosphate-binding domain of DDX24. Knockdown of DDX24 expression in endothelial cells resulted in elevated migration and tube formation. Transcriptomic analysis linked DDX24 to vascular system-related functions. Conclusion: Our results provide a link between DDX24 and vascular malformation and indicate a crucial role for DDX24 in endothelial cell functions; these findings create an opportunity for genetic diagnosis and therapeutic targeting of malformations of vessels to the viscera.
Assuntos
Quilotórax/genética , RNA Helicases DEAD-box/genética , Malformações Vasculares/genética , Vísceras/irrigação sanguínea , Adulto , Sequência de Aminoácidos , Movimento Celular , Células Endoteliais/fisiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Modelos Químicos , Mutação , Linhagem , Conformação ProteicaRESUMO
BACKGROUND: Our previous study showed that overexpression of hepatocyte nuclear factor 4α (HNF4α) could directly promote mesenchymal stem cells (MSCs) to differentiate into hepatocyte-like cells. However, the efficiency of hepatic differentiation remains low. The purpose of our study was to establish an MSC cell line that overexpressed HNF4α and FOXA2 genes to obtain an increased hepatic differentiation efficiency and hepatocyte-like cells with more mature hepatocyte functions. METHODS: Successful establishment of high-level HNF4α and FOXA2 co-overexpression in human induced hepatocyte-like cells (hiHep cells) was verified by flow cytometry, immunofluorescence and RT-PCR. Measurements of albumin (ALB), urea, glucose, indocyanine green (ICG) uptake and release, cytochrome P450 (CYP) activity and gene expression were used to analyze mature hepatic functions of hiHep cells. RESULTS: hiHep cells efficiently express HNF4α and FOXA2 genes and proteins, exhibit typical epithelial morphology and acquire mature hepatocyte-like cell functions, including ALB secretion, urea production, ICG uptake and release, and glycogen storage. hiHep cells can be activated by CYP inducers. The percentage of both ALB and α-1-antitrypsin (AAT)-positive cells was approximately 72.6%. The expression levels of hepatocyte-specific genes (ALB, AAT, and CYP1A1) and liver drug transport-related genes (ABCB1, ABCG2, and SLC22A18) in hiHep cells were significantly higher than those in MSCs-Vector cells. The hiHep cells did not form tumors after subcutaneous xenograft in BALB/c nude mice after 2 months. CONCLUSION: This study provides an accessible, feasible and efficient strategy to generate hiHep cells from MSCs.
Assuntos
Diferenciação Celular , Fator 3-beta Nuclear de Hepatócito/metabolismo , Fator 4 Nuclear de Hepatócito/metabolismo , Hepatócitos/metabolismo , Células-Tronco Mesenquimais/metabolismo , Animais , Biomarcadores/metabolismo , Linhagem Celular , Proliferação de Células , Forma Celular , Feminino , Fator 3-beta Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/genética , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transdução de Sinais , Transfecção , Regulação para CimaRESUMO
BACKGROUND: Many studies have combined sorafenib with transcatheter arterial chemoembolization (TACE) to treat patients with advanced hepatocellular carcinoma (HCC), but the results are disputable. Thus, we conducted this meta-analysis to assess the efficacy and safety of the combination treatment in patients with advanced HCC. METHODS: Clinical data were collected from a computer search of literature published from January 2009 to June 2016 in PubMed, Web of Science, the Cochrane Library, China National Knowledge Infrastructure (CNKI), Wan Fang and the China Science and Technology Journal Database (CSTJ). The final analysis included 14 studies and 1670 patients. The primary endpoints were overall survival (OS), the objective response rate (ORR) and the disease control rate (DCR). RESULTS: The combination group exhibited significantly more improvement than the group treated with TACE alone in ORR (RR =1.62, 95% confidence interval (CI) = 1.34-1.94, p < 0.00001), DCR (RR = 1.43, 95% CI = 1.26-1.62, p < 0.00001), 0.5-year OS (OR = 2.60, 95% CI = 1.57-4.29, p = 0.0002) and 1-year OS (OR = 1.88, 95% CI =1.39-2.53, p < 0.0001). The incidence of adverse events from combination therapy was increased compared to that from treatment with TACE alone, and the most commonly reported adverse events were fatigue, hand-foot skin reaction and diarrhoea, which were bearable. CONCLUSIONS: The meta-analysis indicated that combination therapy is safe and efficient for clinical application.
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Antineoplásicos/administração & dosagem , Carcinoma Hepatocelular/terapia , Quimioembolização Terapêutica/métodos , Neoplasias Hepáticas/terapia , Niacinamida/análogos & derivados , Compostos de Fenilureia/administração & dosagem , Idoso , Idoso de 80 Anos ou mais , Antineoplásicos/uso terapêutico , Terapia Combinada , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Niacinamida/administração & dosagem , Niacinamida/uso terapêutico , Compostos de Fenilureia/uso terapêutico , Sorafenibe , Análise de Sobrevida , Resultado do TratamentoRESUMO
The deregulation of microRNA (miRNA) is frequently associated with a variety of cancers, including hepatocellular carcinoma (HCC). In this study, we investigated the expression and possible role of miR-199a-5p in HCC. The expression of miR-199a-5p was measured by quantitative RT-PCR in HCC. The effect of miR-199a-5p was evaluated by cell viability and colony formation assays in HCC cell lines and tumor cell growth assay in xenograft nude mice. Quantitative real time PCR results showed that miR-199a-5p was down-regulated in 77.9 % (67/86) of HCC tissues compared with adjacent nontumor tissues. MiR-199a-5p mimic reduced cell viability and colony formation by induction of cell arrest in HCC cell lines and inhibited tumor cell growth in xenograft nude mice, but miR-199a-5p inhibitor increased cell viability and colony formation in HCC cell lines and tumor cell growth in xenograft nude mice. Furthermore, CLTC was defined as a potential direct target of miR-199a-5p by MiRanda and TargetScan predictions. The dual-luciferase reporter gene assay results showed that CLTC was a direct target of miR-199a-5p. The use of miR-199a-5p mimic or inhibitor could decrease or increase CLTC protein levels in HCC cell lines. We conclude that the frequently down-regulated miR-199a-5p can regulate CLTC and might function as a tumor suppressor in HCC. Therefore, miR-199a-5p may serve as a useful therapeutic agent for miRNA-based HCC therapy.
Assuntos
Carcinogênese/genética , Carcinoma Hepatocelular/genética , Cadeias Pesadas de Clatrina/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/genética , MicroRNAs/genética , Idoso , Animais , Carcinogênese/metabolismo , Carcinogênese/patologia , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Cadeias Pesadas de Clatrina/metabolismo , Feminino , Genes Reporter , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Luciferases/genética , Luciferases/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Gradação de Tumores , Estadiamento de Neoplasias , Transplante de Neoplasias , Transdução de SinaisRESUMO
OBJECTIVES: To investigate the survival benefit of transarterial chemoembolization (TACE) plus Iodine125 seed implantation (TACE-Iodine125) in hepatitis B-related HCC patients with portal vein tumour thrombus (PVTT) and the underlying prognostic factors. METHODS: A retrospective matched cohort study was performed on consecutive HCC patients with PVTT from January 2011 to June 2014. Seventy patients (TACE-Iodine125 group) who underwent TACE-Iodine125 were compared with a historical case-matched control group of 140 patients (TACE group) who received TACE alone. The survival of patients and the underlying prognostic factors were analysed. RESULTS: The median survival times of the TACE-Iodine125 and TACE groups were 11.0 and 7.5 months, respectively (p < 0.001). The survival probability at 12, 24, and 36 months was 50 %, 14.5 %, and 14.5 % vs. 25 %, 9 %, and 5 % in the TACE-Iodine125 and TACE groups, respectively (p < 0.001). The PVTT responders had better survival than the PVTT non-responders (p < 0.001). For the PVTT non-responders, there were no differences in the survival curves between the groups (p = 0.353). Multivariate analysis showed that type III PVTT (p < 0.001) and APS (p < 0.001) were independent predictors of poor prognosis. In contrast, the treatment modality of TACE-Iodine125 (p < 0.001) and PVTT response (p = 0.001) were favourable prognostic features. CONCLUSIONS: TACE combined with Iodine125 seed implantation may be a good choice for selected HB-HCC patients with PVTT. KEY POINTS: ⢠TACE-Iodine125 was more effective than TACE for patients with HCC-PVTT. ⢠The TACE-Iodine125 procedure was safe. ⢠TACE-Iodine125 was conditional for patients with HCC-PVTT. ⢠TACE-Iodine125 resulted in a better PVTT response compared to TACE alone. ⢠A good PVTT response is a favourable prognostic factor.
Assuntos
Braquiterapia/métodos , Carcinoma Hepatocelular/terapia , Quimioembolização Terapêutica/métodos , Hepatite B/complicações , Radioisótopos do Iodo/uso terapêutico , Neoplasias Hepáticas/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Hepatocelular/complicações , Estudos de Coortes , Feminino , Humanos , Neoplasias Hepáticas/complicações , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Análise de Sobrevida , Resultado do Tratamento , Adulto JovemRESUMO
OBJECTIVE: To examine the feasibility and efficacy of using superparamagnetic iron oxide nanoparticles coated with polyethylene glycol-grafted polyethylenimine (PEG-g-PEI-SPION) as a carrier for gene delivery into human adipose derived mesenchymal stem cells (hADMSCs) and in vitro cellular magnetic resonance imaging (MRI). METHODS: PEG-g-PEI-SPION was synthesized as previously reported. Gel electrophoresis was performed to assess the pDNA condensation capacity of PEG-g-PEI-SPION. The particle size and zeta potential of PEG-g-PEI-SPION/pDNA complexes were determined by dynamic light scattering. Cytotoxicity of PEG-g-PEI-SPION was evaluated by CCK-8 assay with hADMSCs. Gene transfection efficiency of PEG-g-PEI-SPION in hADMSCs was quantified by flow cytometry. The cellular internalization of PEG-g-PEI-SPION/pDNA nanocomplexes was studied by confocal laser scanning microscopy and Prussian blue staining. MRI function of PEG-g-PEI-SPION was studied by in vitro cellular MRI scanning. RESULTS: PEG-g-PEI-SPION condensed pDNA to form stable complexes of 80-100 nm in diameter and showed low cytotoxicity in hADMSCs. At the optimal N/P ratio of 20, PEG-g-PEI-SPION/pDNA obtained the highest transfection efficiency of 22.8% ± 3.6% in hADMSCs. And it was higher than that obtained with lipofectamine 11.2% ± 2.6% (P < 0.05). Furthermore, hADMSCs labeled with PEG-g-PEI-SPION showed sensitive low signal intensity on MRI T2-weighted images in vitro. CONCLUSION: PEG-g-PEI-SPION is an efficient and MRI-visible nano-vector for gene delivery into hADMSCs.
Assuntos
Técnicas de Transferência de Genes , Células-Tronco Mesenquimais/citologia , Transfecção , Tecido Adiposo/citologia , Células Cultivadas , Vetores Genéticos , Humanos , Imageamento por Ressonância Magnética/métodos , NanopartículasRESUMO
OBJECTIVE: Endovenous microwave ablation (EMA) is a recently developed thermal ablation technique used in the treatment of lower limb varicose veins. However, its efficacy and safety have been largely understudied. In the present study, we sought to explore the clinical results of EMA and radiofrequency ablation (RFA) in treating lower limb varicose veins. METHODS: Patients who underwent EMA (n = 65) or RFA (n = 46) at our institute from September 2018 to September 2020 were included in this retrospective investigation. The clinical results and complications were evaluated at 1, 3, 6, and 12 months after the procedure. The effects on disease severity and quality of life were evaluated using the venous clinical severity score and chronic venous insufficiency questionnaire (CIVIQ). RESULTS: The technical success rate was 100% for both experimental groups. Although the operative time between the two groups was comparable, the EMA technique was associated with lower direct costs (P < .001), although also with prolonged hospitalization (P < .001). We found that the use of EMA correlated with more pain at 48 hours postoperatively. Except for the visual analog scale scores, no statistically significant variations were observed in the occurrence of postoperative complications within the first 48 hours postoperatively between the EMA and RFA groups, including paresthesia, ecchymosis, induration, and phlebitis (P > .05). At 4 weeks postoperatively, significantly less pigmentation was observed in the RFA group than in the EMA group (13.04% vs 32.31%; P = .020). However, the pigmentation had resolved in all patients by 12 months postoperatively. The two groups had a reduction in the venous clinical severity scores and an increase in the CIVIQ scores after the procedure. However, the CIVIQ scores within the RFA group had increased more than had those within the EMA group (P < .05). No significant differences were found in recurrence between the two groups (EMA group, 1.54%; RFA group, 2.17%; P = .804). CONCLUSIONS: Both ablation techniques are safe and effective. RFA is associated with relatively higher treatment costs but shorter hospitalization and better quality of life improvement.
Assuntos
Ablação por Cateter , Terapia a Laser , Ablação por Radiofrequência , Varizes , Insuficiência Venosa , Humanos , Varizes/diagnóstico por imagem , Varizes/cirurgia , Qualidade de Vida , Estudos Retrospectivos , Micro-Ondas/efeitos adversos , Insuficiência Venosa/diagnóstico por imagem , Insuficiência Venosa/cirurgia , Ablação por Radiofrequência/efeitos adversos , Veia Safena/cirurgia , Resultado do Tratamento , Ablação por Cateter/efeitos adversos , Ablação por Cateter/métodos , Terapia a Laser/métodosRESUMO
BACKGROUND: Acute hepatitis is a progressive inflammatory disorder that can lead to liver failure. Endothelial permeability is the vital pathophysiological change involved in infiltrating inflammatory factors. DDX24 has been implicated in immune signaling. However, the precise role of DDX24 in immune-mediated hepatitis remains unclear. Here, we investigate the phenotype of endothelium-targeted Ddx24 conditional knockout mice with Concanavalin A (ConA)-induced hepatitis. METHODS: Mice with homozygous endothelium-targeted Ddx24 conditional knockout (Ddx24flox/flox; Cdh5-Cre+) were established using the CRISPR/Cas9 mediated Cre-loxP system. We investigated the biological functions of endothelial cells derived from transgenic mice and explored the effects of Ddx24 in mice with ConA-induced hepatitis in vivo. The mass spectrometry was performed to identify the differentially expressed proteins in liver tissues of transgenic mice. RESULT: We successfully established mice with endothelium-targeted Ddx24 conditional knockout. The results showed migration and tube formation potentials of murine aortic endothelial cells with DDX24 silencing were significantly promoted. No differences were observed between Ddx24flox/flox; Cdh5-Cre+ and control regarding body weight and length, pathological tissue change and embryogenesis. We demonstrated Ddx24flox/flox; Cdh5-Cre+ exhibited exacerbation of ConA-induced hepatitis by up-regulating TNF-α and IFN-γ. Furthermore, endothelium-targeted Ddx24 conditional knockout caused vascular hyper-permeability in ConA-injected mice by down-regulating vascular integrity-associated proteins. Mechanistically, we identified Ddx24 might regulate immune-mediated hepatitis by inflammation-related permeable barrier pathways. CONCLUSION: These findings prove that endothelium-targeted Ddx24 conditional knockout exacerbates ConA-induced hepatitis in mice because of vascular hyper-permeability. The findings indicate a crucial role of DDX24 in regulating immune-mediated hepatitis, suggesting DDX24 as a potential therapeutic target in the disorder.