RESUMO
The activation of hepatic stellate cells (HSCs) plays a key role in the occurrence of liver fibrosisï¼and promoting the apoptosis of activated HSCs or reducing the number of activated HSCs can reverse the development of liver fibrosis. In our previous studies, we have demonstrated that the CCAAT/enhancer binding protein α (C/EBP-α) played an important role in promoting the apoptosis of activated HSCs, thereby exerting an anti-liver fibrosis effect. Unlike apoptosis, autophagy, as a caspase-independent programmed cell death, can promptly remove the abnormal accumulation of substances or damaged organelles in cells and play a key role in regulating the homeostasis of intracellular environment. However, it is still unclear whether C/EBP-α participates in the occurrence of autophagy in HSCs. Therefore, in this study, we firstly used the methods of Western blot and immunofluorescence to characterize the consequence of C/EBP-α overexpression on the expression of proteins LC3B, P62, ATG5 and Beclin1 which were related to autophagy in HSCs. Subsequently, we performed Western blot and site-directed mutagenesis methods to clarify the type and related mechanism of autophagy which was induced by C/EBP-α. Here we show that C/EBP-α promotes the occurrence of autophagy in HSCs and the autophagy induced by C/EBP-α belongs to mitophagy. The stability of C/EBP-α protein regulates the level of autophagy in HSCs. In addition, acetylation of C/EBP-α also regulates the occurrence of autophagy in HSCs. Acetylation of lysine at positions K298, K302 and K326 of C/EBP-α promotes its binding to Beclin1. In conclusion, our study uncovers the role of C/EBP-α in regulating autophagy in HSCs, thereby providing a new strategy for clinical treatment of liver fibrosis.
Assuntos
Autofagia/fisiologia , Proteína Beclina-1/metabolismo , Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Células Estreladas do Fígado/citologia , Acetilação , Animais , Homeostase/fisiologia , Cirrose Hepática/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , RatosRESUMO
AIMS: To analyze the clinicopathological characteristics of poorly differentiated chordomas (PDCs) with SMARCB1/INI1 loss in children. MATERIALS: Four cases of PDCs were included in the study. METHODS: Immunohistochemistry was performed with respect to brachyury, Glut-1, keratin 18, keratin 19, INI1, vimentin, S-100, CK, EMA, GFAP, etc. Fluorescence in situ hybridization (FISH) was performed for SMARCB1/INI1 from 3 patients. RESULTS: Histologically, contrary to typical histologic features for conventional chordomas, 4 tumors were composed of ovoid or atypical fusiform cells. Sporadic physaliphorous cells were evident. Tumor cells had large vacuoles in the cytoplasm that were even remarkable on the imprint cytology slide. By immunohistochemistry, each case revealed loss of SMARCB1/INI1 expression and nuclear expression of brachyury. Glut-1, keratin 18, keratin 19, CK, EMA, and vimentin were positive in these PDCs. Except for 1 patient who had not yet completed FISH, the other 3 cases demonstrated the loss of SMARCB1/INI1 gene by fluorescence in situ hybridization. CONCLUSION: Poorly differentiated SMARCB1/INI1-negative chordoma is a unique subset of chordoma representing a clinically, histopathologically, and molecularly distinct entity with rapid progression and poor prognosis which should not be confused with conventional chordomas. Sporadic physaliphorous cells (tumor cells with large vacuoles in the cytoplasm) provided important diagnostic clues of PDCs. Combination use of characteristic markers of notochord cells (brachyury, Glut-1, keratin 18, and keratin 19) along with INI1 were effective diagnostic tools.
Assuntos
Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/genética , Cordoma/diagnóstico , Cordoma/genética , Proteína SMARCB1/genética , Adolescente , Criança , Pré-Escolar , Feminino , Humanos , Hibridização in Situ Fluorescente , Imageamento por Ressonância Magnética , Masculino , Adulto JovemRESUMO
Dysregulation of iron metabolism and aberrant iron deposition has been associated with multiple sclerosis. However, the factors that contribute to this pathological state remain to be understood. In this study, human multiple sclerosis and mice brain samples were analyzed through mass spectrometry as well as histological and immunoblot techniques, which demonstrated that iron deposition is associated with increased levels of nuclear prelamin A recognition factor (NARF). NARF is a protein associated with the mitochondria which has also been linked to mitochondrial defects in multiple sclerosis. We report NARF to be associated in multiple sclerosis pathology and aberrant iron deposition.
Assuntos
Encéfalo/metabolismo , Ferro/metabolismo , Lamina Tipo A/metabolismo , Esclerose Múltipla/metabolismo , Encéfalo/patologia , Células HEK293 , Humanos , Esclerose Múltipla/patologia , Proteínas Nucleares/metabolismoRESUMO
We used the concept of bioisosteres to design and synthesize a novel series of dasatinib derivatives for the treatment of leukemia. Unfortunately, most of the dasatinib derivatives did not show appreciable inhibition against leukemia cell lines K562 and HL60. However, acrylamide compound 2c had comparable inhibitory activity with dasatinib against K562 cells (IC50â¯=â¯0.039â¯nM vs. 0.069â¯nM). And amide compound 2a and acrylamide compound 2c also had comparable inhibitory activity with dasatinib against the leukemia cell line HL60 (IC50â¯=â¯0.25â¯nM and 0.26â¯nM vs. 0.11â¯nM). Against the leukemia progenitor cell line KG1a, triazole compounds 15a and 15d-15f and oxadiazole compounds 24a-24d were more potent than dasatinib. In particular, the hydroxyl compounds 15a and 24a were about 64 and 180 fold more potent than dasatinib against KG1a cells (IC50â¯=â¯0.14⯵M and 0.05⯵M vs. 8.98⯵M). Compounds 15a and 24a also inhibited colony formation in MCF-7 cells and inhibited cell migration in the cell wound scratch assay in B16BL6 cells. Moreover, hydroxyl compounds 15a and 24a had low toxicity in vivo.
Assuntos
Antineoplásicos/farmacologia , Dasatinibe/análogos & derivados , Dasatinibe/farmacologia , Leucemia/tratamento farmacológico , Células-Tronco Neoplásicas/efeitos dos fármacos , Animais , Antineoplásicos/síntese química , Antineoplásicos/toxicidade , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Dasatinibe/síntese química , Dasatinibe/toxicidade , Desenho de Fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Camundongos , Oxidiazóis/síntese química , Oxidiazóis/farmacologia , Oxidiazóis/toxicidade , Triazóis/síntese química , Triazóis/farmacologia , Triazóis/toxicidadeAssuntos
Carcinoma de Células Renais , Neoplasias Renais , Neoplasias de Células Epitelioides Perivasculares , Feminino , Humanos , Criança , Neoplasias de Células Epitelioides Perivasculares/genética , Neoplasias de Células Epitelioides Perivasculares/patologia , Carcinoma de Células Renais/patologia , Neoplasias Renais/genética , Neoplasias Renais/patologia , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/genética , Rim/patologia , Translocação Genética , Cromossomos Humanos X/genética , Biomarcadores TumoraisRESUMO
Glycosylation analysis of proteins from biological sources utilizing mass spectrometry based approaches is challenging due to the relatively low abundance of glycopeptides, the structural diversity of glycans, and the coexisting matrices. In this study, a customized dextran-bonded silica-based stationary phase was introduced for selective enrichment of glycopeptides and glycans from complex biological samples. This material has exhibited superior selectivity and broader glycosylation site coverage over commercial Sepharose in glycoproteomic evaluation. Additionally, the glycomic analysis of fetuin, α1 -acid glycoprotein, and human serum N-glycome also indicated the relatively higher sensitivity, selectivity, and glycoform coverage of dextran-bonded silica than that of Sepharose and porous graphitized carbon. Therefore, the dextran-bonded silica is expected to make contributions in the fields of glycoproteomics and glycomics.
Assuntos
Dextranos/química , Glicopeptídeos/química , Polissacarídeos/química , Adulto , Animais , Carbono/química , Bovinos , Fetuínas/química , Glicômica , Glicosilação , Humanos , Interações Hidrofóbicas e Hidrofílicas , Masculino , Espectrometria de Massas , Peptídeos/química , Proteômica , Sefarose/química , Soro/química , Dióxido de Silício/químicaRESUMO
Deimination refers to conversion of protein-bound arginine into citrulline. An mRNA carrier, RNA binding export factor (REF), present on mitochondria undergoes loss of deimination with impaired ATP5b mRNA transport in ND4 mice (model of multiple sclerosis) compared with the controls. We present evidence of (1) reduced ATP5b mRNA binding strength of non-deiminated REF compared with deiminated REF, (2) impaired ATP5b mRNA transport in ND4 mice and (3) reduced mitochondrial ATP synthase activity on inhibition of deimination in PC12 cells. Impaired deimination of REF and defect in mitochondrial mRNA transport are critical factors in mitochondrial dysfunction in ND4 mice.
Assuntos
ATPases Mitocondriais Próton-Translocadoras/genética , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Esclerose Múltipla/genética , Esclerose Múltipla/metabolismo , RNA Mensageiro/metabolismo , Complexos de ATP Sintetase/metabolismo , Trifosfato de Adenosina/biossíntese , Animais , Linhagem Celular , Citrulina/análise , Modelos Animais de Doenças , Ativação Enzimática , Expressão Gênica , Camundongos , Camundongos Transgênicos , Mitocôndrias/metabolismo , Proteínas Nucleares/metabolismo , Ligação Proteica , Transporte Proteico , Transporte de RNA , Proteínas de Ligação a RNA/metabolismo , Ratos , Fatores de Transcrição/metabolismoRESUMO
Aims/Background: Insulinoma is an extremely rare condition in pediatric patients. This study aims to examine the pathological and clinical characteristics of pediatric insulinoma. Methods: A retrospective, single-center study was conducted involving five pediatric patients diagnosed with insulinoma. The study involved evaluating the postoperative status of the patients during follow-up and analyzing their clinical manifestations, diagnostic work-up, pathological findings, and therapeutic approaches. Results: The study cohort comprised four males and one female, aged between 4 and 9 years. Common symptoms included dizziness and fatigue. The insulinomas were located in various parts of the pancreas: two in the head, one in the neck, one in the body, and one in the tail. After undergoing subtotal pancreatectomy, four patients experienced no side effects during a follow-up period of 41 to 153 months. One patient, who underwent an incomplete pancreatic resection, required ongoing postoperative treatment with 150 mg Creon due to pancreatic enzyme deficiency. Postoperative pathological results indicated that all cases were low-grade neuroendocrine tumours, classified as grade 1 (G1) or grade 2 (G2). Two cases exhibited capsule invasion, and one case showed microvascular invasion. Despite these invasions, no recurrences or metastases have been observed to date. Conclusion: Surgical resection is a viable treatment option for pediatric insulinoma, yielding a favorable prognosis. The presence of capsular and microvascular invasions does not seem to affect the overall prognosis in these cases.
Assuntos
Insulinoma , Pancreatectomia , Neoplasias Pancreáticas , Humanos , Insulinoma/cirurgia , Insulinoma/patologia , Insulinoma/diagnóstico , Masculino , Feminino , Criança , Estudos Retrospectivos , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/cirurgia , Pré-Escolar , Pancreatectomia/métodosRESUMO
Bacterial panicle blight, bacterial leaf streak, and bacterial brown stripe are common bacterial diseases in rice that represent global threats to stable rice yields. In this study, we used the rice variety HZ, Nekken and their 120 RIL population as experimental materials. Phenotypes of the parents and RILs were quantitatively analyzed after inoculation with Burkholderia glumae, Xanthomonas oryzae pv. oryzicola, and Acidovorax avenae subsp. avenae. Genetic SNP maps were also constructed and used for QTL mapping of the quantitative traits. We located 40 QTL loci on 12 chromosomes. The analysis of disease resistance-related candidate genes in the QTL regions with high LOD value on chromosomes 1, 3, 4, and 12 revealed differential expression before and after treatment, suggesting that the identified genes mediated the variable disease resistance profiles of Huazhan and Nekken2. These results provide an important foundation for cloning bacterial-resistant QTLs of panicle blight, leaf streak, and brown stripe in rice.
RESUMO
CCAAT enhancer binding protein-α (C/EBP-α) is a transcript factor that regulates adipocyte differentiation and induces apoptosis in hepatic stellate cells (HSCs) in vivo and in vitro. However, the effect of C/EBP-α on hepatocytes in vivo remains unknown. This study investigated whether C/EBP-α exerts different apoptotic effects on hepatocytes and HSCs in vitro and in vivo. An adenovirus vector-expressing C/EBP-α gene was constructed, and a rat hepatic stellate cell lines (HSC-T6) and hepatocytes were transfected. A CCl(4)-induced liver fibrosis model in mice was also utilized. C/EBP-α induced apoptosis in hepatocytes and HSCs, but a significant difference between these cell types was observed in vitro. The mitochondrial pathway was involved in the apoptotic process and was predominant in HSC-T6 apoptosis. In the CCl(4)-induced mice liver fibrosis model, the administration of Ad-C/EBP-α decreased extracellular matrix deposition, including collagen and hydroxyproline content, and γ-GT levels, a marker of liver damage, were reduced significantly. Immunohistochemistry and TUNEL assay results showed an increase of apoptosis in HSCs, but hepatocytes were less affected. C/EBP-α induced differential apoptotic effects in hepatocytes and HSCs in vitro and in vivo. This differential effect could be a potential target for the treatment of hepatic fibrosis with little hepatic toxicity.
Assuntos
Apoptose , Proteína alfa Estimuladora de Ligação a CCAAT/fisiologia , Células Estreladas do Fígado/fisiologia , Hepatócitos/fisiologia , Cirrose Hepática/metabolismo , Actinas/metabolismo , Animais , Proteína alfa Estimuladora de Ligação a CCAAT/biossíntese , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Tetracloreto de Carbono , Caspases/genética , Caspases/metabolismo , Células Cultivadas , Colágeno/metabolismo , Fragmentação do DNA , Matriz Extracelular/metabolismo , Expressão Gênica , Células Estreladas do Fígado/metabolismo , Hepatócitos/metabolismo , Hidroxiprolina/metabolismo , Cirrose Hepática/induzido quimicamente , Cirrose Hepática/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Ratos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , gama-Glutamiltransferase/metabolismoRESUMO
To investigate the diagnostic utility of immunohistochemistry for paired-like homeobox 2B (PHOX2B) expression in neuroblastomas (NBs) and tumors that mimic them, tissue samples (n = 229) from 157 cases of NB, 210 central nervous system tumors, and 170 extracranial non-NB solid tumors (n = 170) were immunostained for PHOX2B. Additionally, PHOX2B expression in 67 body fluid cytology specimens was analyzed. In tissue specimens, PHOX2B expression was positive in NBs, pheochromocytomas, and paragangliomas but negative in all of the other tumors evaluated. PHOX2B was detected by immunohistochemistry in 5 NB cytology specimens; all of the others were negative. These results suggest that PHOX2B may be a sensitive and specific immunohistochemical marker for the pathological diagnosis and differential diagnosis of NB in both tissue and cytology specimens.
Assuntos
Biomarcadores Tumorais/metabolismo , Proteínas de Homeodomínio/metabolismo , Neuroblastoma/diagnóstico , Fatores de Transcrição/metabolismo , Biomarcadores Tumorais/análise , Proteínas de Homeodomínio/análise , Humanos , Imuno-Histoquímica , Fatores de Transcrição/análiseRESUMO
Colorectal cancer is one of the top leading causes of cancer mortality worldwide, especially in China. However, most of the current treatments are invasive and can only be applied to very few cancers. The earlier a malignant tumor is diagnosed, the higher the patient's survival rate. In this study, we proposed a computational framework to identify highly-reliable and easierly-detectable biomarkers capable of secreting into blood, urine and saliva by integrating transcriptomics and proteomics data at the system biology level. First, a large number of transcriptome data were processed to identify candidate biomarkers for colorectal cancer. Second, three classified models are constructed to predict biomarkers for colorectal cancer capable of secreting into blood, urine and saliva, which are effective disease diagnosis media to facilitate clinical screening. Then biological functions and molecular mechanisms of the candidate biomarkers of colorectal cancer are inferred utilizing multi-source biological knowledge and literature mining. Furthermore, the classification power of different combinations of candidate biomarkers is verified by machine learning models. In addition, the targeted drugs of the predicted biomarkers are further analyzed to provide assistance for clinical treatment of colorectal cancer. In this paper, our proposed computational model not only provides the effective candidate biomarkers ESM1, CTHRC1, AZGP1 for colorectal cancer capable of secreting into blood, urine and saliva, but also helps to understand the molecular mechanism of colorectal cancer. This computational framework can span the huge gap between transcriptome and proteomics, which can easily be applied to the biomarker research for other types of tumor.
Assuntos
Biomarcadores Tumorais/análise , Neoplasias Colorretais/diagnóstico , Biologia Computacional , Proteínas de Neoplasias/análise , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/genética , Neoplasias Colorretais/metabolismo , Bases de Dados de Proteínas , Humanos , Aprendizado de Máquina , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , ProteômicaRESUMO
Neurons from the adult central nervous system (CNS) demonstrate limited mRNA transport and localized protein synthesis versus developing neurons, correlating with lower regenerative capacity. We found that deimination (posttranslational conversion of protein-bound arginine into citrulline) undergoes upregulation during early neuronal development while declining to a low basal level in adults. This modification is associated with neuronal arborization from amphibians to mammals. The mRNA-binding proteins (ANP32a, REF), deiminated in neurons, have been implicated in local protein synthesis. Overexpression of the deiminating cytosolic enzyme peptidyl arginine deiminase 2 in nervous systems results in increased neuronal transport and neurite outgrowth. We further demonstrate that enriching deiminated proteins rescues transport deficiencies both in primary neurons and mouse optic nerve even in the presence of pharmacological transport blockers. We conclude that deimination promotes neuronal outgrowth via enhanced transport and local protein synthesis and represents a new avenue for neuronal regeneration in the adult CNS.
Assuntos
Reprogramação Celular , Iminas/metabolismo , Regeneração Nervosa , Neurônios/metabolismo , Sequência de Aminoácidos , Animais , Polaridade Celular , Proliferação de Células , Modelos Animais de Doenças , Camundongos , Células PC12 , Desiminases de Arginina em Proteínas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RatosRESUMO
Reversal of activated hepatic stellate cells (HSCs) to a quiescent state and apoptosis of activated HSCs are key elements in the reversion of hepatic fibrosis. CCAAT/enhancer binding protein α (C/EBP-α) has been shown to inhibit HSC activation and promote its apoptosis. This study aims to investigate how C/EBP-α acetylation affects the fate of activated HSCs. Effects of a histone deacetylation inhibitor trichostatin A (TSA) on HSC activation were evaluated in a mouse model of liver fibrosis caused by carbon tetrachloride (CCl4) intoxication. TSA was found to ameliorate CCl4-induced hepatic fibrosis and improve liver function through increasing the protein level and enhancing C/EBP-α acetylation in the mouse liver. C/EBP-α acetylation was determined in HSC lines in the presence or absence of TSA, and the lysine residue K276 was identified as a main acetylation site in C/EBP-α protein. C/EBP-α acetylation increased its stability and protein level, and inhibited HSC activation. The present study demonstrated that C/EBP-α acetylation increases the protein level by inhibiting its ubiquitination-mediated degradation, and may be involved in the fate of activated HSCs. Use of TSA may confer an option in minimizing hepatic fibrosis by suppressing HSC activation, a key process in the initiation and progression of hepatic fibrosis.
Assuntos
Proteína alfa Estimuladora de Ligação a CCAAT/metabolismo , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Ácidos Hidroxâmicos/farmacologia , Acetilação , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Sítios de Ligação , Biomarcadores , Proteína alfa Estimuladora de Ligação a CCAAT/genética , Tetracloreto de Carbono/efeitos adversos , Linhagem Celular , Expressão Gênica , Células Estreladas do Fígado/patologia , Humanos , Imuno-Histoquímica , Cirrose Hepática/etiologia , Cirrose Hepática/metabolismo , Cirrose Hepática/patologia , Masculino , Camundongos , Mutação , Ligação Proteica , Estabilidade Proteica , Ratos , UbiquitinaçãoRESUMO
CCAAT enhancer binding proteinα (C/EBPα) is a transcription factor expressed only in certain tissues, including the liver. It has been previously demonstrated that C/EBPα may induce apoptosis in hepatic stellate cells (HSCs), raising the question of whether acetylation of C/EBPα is associated with HSCs, and the potential associated mechanism. A total of three histone deacetylase inhibitors (HDACIs), including trichostatin A (TSA), suberoylanilide hydroxamic acid and nicotinamide, were selected to determine whether acetylation affects C/EBPα expression. A Cell Counting Kit8 assay was used to determine the rate of proliferation inhibition following treatment with varying doses of the three HDACIs in HSCT6 and BRL3A cells. Western blot analysis was used to examine Caspase3, 8, 9, and 12 levels in HSCT6 cells treated with adenoviralC/EBPα and/or TSA. Following treatment with TSA, a combination of reverse transcriptionquantitative polymerase chain reaction and western blot analyses was used to determine the inherent C/EBPα mRNA and protein levels in HSCT6 cells at 0, 1, 2, 4, 8, 12, 24, 36 and 48 h. Nuclear and cytoplasmic proteins were extracted to examine C/EBPα distribution. Coimmunoprecipitation analysis was used to examine the lysine acetylation of C/EBPα. It was observed that TSA inhibited the proliferation of HSCT6 cells to a greater extent compared with BRL3A cells, following treatment with the three HDACIs. TSA induced apoptosis in HSCT6 cells and enhanced the expression of C/EBPα. Following treatment of HSCT6 cells with TSA, inherent C/EBPα expression increased in a timedependent manner, and its lysine acetylation simultaneously increased. Therefore, the results of the present study suggested that TSA may increase C/EBPα expression by increasing its lysine acetylation in HSCs.