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1.
Intern Med ; 63(8): 1093-1097, 2024 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-37661447

RESUMO

A male patient in his 70s with recurrent hepatocellular carcinoma (HCC) after surgery received atezolizumab plus bevacizumab (Atezo+Bev) therapy. Initial computed tomography (CT) revealed tumor growth along with an increase in tumor markers, and contrast-enhanced ultrasonography (CEUS) showed multiple round avascular areas within the nodules with an appearance similar to a slice of Swiss cheese. Continuation of immunotherapy with consideration of the potential for pseudoprogression produced a dramatic response. Although it is difficult to distinguish between true progression and pseudoprogression, the Swiss cheese-like appearance on CEUS may be important for the early diagnosis of pseudoprogression.


Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Masculino , Carcinoma Hepatocelular/diagnóstico por imagem , Carcinoma Hepatocelular/terapia , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/terapia , Neoplasias Hepáticas/patologia , Meios de Contraste , Ultrassonografia/métodos , Imunoterapia
2.
J Gastroenterol Hepatol ; 38(10): 1847-1854, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37646384

RESUMO

BACKGROUND AND AIM: There have been several reports that some probiotics improve non-alcoholic fatty liver disease (NAFLD); however, many studies have involved cocktail therapies. We evaluated whether heat-killed Lactobacillus brevis SBL88 (L. brevis SBL88) monotherapy improves the clinical features of NAFLD. METHODS: The NAFLD model was induced in mice fed a high-fat diet (HFD) (HFD mice) or HFD + 1% heat-killed L. brevis SBL88 (SBL mice) for 16 weeks. Histopathological liver findings were analyzed. To evaluate the gut microbiota, a modified terminal restriction fragment length polymorphism analysis of the feces was performed. RNA sequencing in the liver was performed with Ion Proton™. To investigate the direct effects of heat-killed L. brevis SBL88, an in vitro study was performed. RESULTS: Histopathological findings revealed that fat droplets in the liver were significantly reduced in SBL mice; however, terminal restriction fragment length polymorphism did not show alterations in the gut microbiota between HFD mice and SBL mice. RNA sequencing and pathway analysis revealed that the regulation of lipid and insulin metabolism was affected. The mRNA expression of insulin receptor substrate 2 (IRS-2) was significantly higher in SBL mice, whereas the expression of IRS-1 was not significantly different. Phospho-IRS-2 expression was also significantly increased in SBL mice. In addition, an in vitro study revealed significant alterations in IRS-2 and forkhead box protein O1 expression levels. CONCLUSION: SBL mice exhibited partially improved selective hepatic insulin resistance. Our data suggest that heat-killed L. brevis SBL88 could attenuate the clinical features of NAFLD that are not mediated by alterations in the gut microbiota.


Assuntos
Microbioma Gastrointestinal , Resistência à Insulina , Levilactobacillus brevis , Hepatopatia Gordurosa não Alcoólica , Animais , Camundongos , Hepatopatia Gordurosa não Alcoólica/terapia , Hepatopatia Gordurosa não Alcoólica/tratamento farmacológico , Resistência à Insulina/genética , Microbioma Gastrointestinal/genética , Temperatura Alta , Fígado/patologia , Dieta Hiperlipídica/efeitos adversos , Camundongos Endogâmicos C57BL
3.
Intern Med ; 61(23): 3497-3502, 2022 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-35491133

RESUMO

A 63-year-old man with hepatitis C was treated with atezolizumab plus bevacizumab for unresectable diffuse hepatocellular carcinoma (HCC). After four cycles of atezolizumab plus bevacizumab, the diffuse HCC markedly shrank; however, he complained of general fatigue, loss of appetite, and slight loss of muscle strength in the lower legs. He was diagnosed with isolated adrenocorticotropic hormone deficiency (IAD), hypothyroidism, and myopathy, suggesting multisystem immune-related adverse events (irAEs). After administration of hydrocortisone, the clinical symptoms rapidly disappeared. Patients with multisystem irAEs can have favorable outcomes; thus, to continue immune-checkpoint inhibitors therapy, a correct diagnosis and management of multisystem irAEs are important.


Assuntos
Carcinoma Hepatocelular , Hepatite C , Neoplasias Hepáticas , Masculino , Humanos , Pessoa de Meia-Idade , Carcinoma Hepatocelular/tratamento farmacológico , Bevacizumab/efeitos adversos , Neoplasias Hepáticas/tratamento farmacológico
4.
Cancer Sci ; 112(2): 563-574, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33211385

RESUMO

Copy number alterations detected by comparative genomic hybridization (CGH) can lead to the identification of novel cancer-related genes. We analyzed chromosomal aberrations in a set of 100 human primary colorectal cancers (CRCs) using CGH and found a solute carrier (SLC) 7A1 gene, which encodes cationic amino acid transporter 1 (CAT1) with 14 putative transmembrane domains, in a chromosome region (13q12.3) with a high frequency of gene amplifications. SLC7A1/CAT1 is a transporter responsible for the uptake of cationic amino acids (arginine, lysine, and ornithine) essential for cellular growth. Microarray and PCR analyses have revealed that mRNA transcribed from CAT1 is overexpressed in more than 70% of human CRC samples, and RNA interference-mediated knockdown of CAT1 inhibited the cell growth of CRCs. Rats were immunized with rat hepatoma cells expressing CAT1 tagged with green fluorescent protein (GFP), and rat splenocytes were fused with mouse myeloma cells. Five rat monoclonal antibodies (mAbs) (CA1 ~ CA5) reacting with HEK293 cells expressing CAT1-GFP in a GFP expression-dependent manner were selected from established hybridoma clones. Novel anti-CAT1 mAbs selectively reacted with human CRC tumor tissues compared with adjacent normal tissues according to immuno-histochemical staining and bound strongly to numerous human cancer cell lines by flow cytometry. Anti-CAT1 mAbs exhibited internalization activity, antibody-dependent cellular cytotoxicity, and migration inhibition activity against CRC cell lines. Furthermore, CA2 inhibited the in vivo growth of human HT29 and SW-C4 CRC tumors in nude mice. This study suggested CAT1 to be a promising target for mAb therapy against CRCs.


Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Transportador 1 de Aminoácidos Catiônicos/antagonistas & inibidores , Neoplasias Colorretais/genética , Animais , Transportador 1 de Aminoácidos Catiônicos/genética , Linhagem Celular Tumoral , Amplificação de Genes , Xenoenxertos , Humanos , Camundongos , Camundongos Nus , Ratos
5.
J Gastroenterol Hepatol ; 35(6): 1042-1048, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31752049

RESUMO

BACKGROUND AND AIM: Because of their survival benefits, immune checkpoint inhibitors (ICIs) are widely administered to patients with various advanced-stage malignancies. During ICI treatment, drug-induced liver injury (DILI) occasionally occurs. In particular, hepatic immune-related adverse events (irAEs) are rare but serious and fatal. In patients with hepatic irAEs, immediate steroid treatment is generally recommended; however, the risk factors for ICI-associated DILI remain unknown. In the present study, we identified a risk factor for ICI-associated DILI. METHODS: We retrospectively analyzed 135 patients treated with anti-programmed cell death-1 (PD-1) antibodies, such as nivolumab and pembrolizumab, at Asahikawa Medical University Hospital. We investigated grade ≥ 2 hepatotoxic AEs during anti-PD-1 therapy, and PD-1 inhibitor-associated DILI was then diagnosed according to the Digestive Disease Week Japan (DDW-J) 2004 scale. The risk factors for PD-1 inhibitor-associated DILI were identified by Cox hazard analysis. RESULTS: Thirty-six patients developed grade ≥ 2 hepatic AEs during anti-PD-1 therapy. Among them, eight patients were diagnosed with PD-1 inhibitor-associated DILI based on the DDW-J 2004 scale. Cox hazard analysis revealed that non-alcoholic fatty liver disease (NAFLD) was a risk factor for PD-1 inhibitor-associated DILI. In addition, we revealed that the outcomes of patients with the DDW-J 2004 score = 3 were improved without steroid treatment. CONCLUSIONS: NAFLD is a potential risk factor for PD-1 inhibitor-associated DILI based on the DDW-J 2004 scale. The DDW-J 2004 scale might be useful for determining whether steroid treatment is required in patients with PD-1 inhibitor-associated DILI.


Assuntos
Anticorpos Monoclonais Humanizados/efeitos adversos , Anticorpos Monoclonais/efeitos adversos , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Neoplasias/tratamento farmacológico , Nivolumabe/efeitos adversos , Hepatopatia Gordurosa não Alcoólica , Receptor de Morte Celular Programada 1/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Monoclonais/administração & dosagem , Anticorpos Monoclonais Humanizados/administração & dosagem , Feminino , Glucocorticoides/administração & dosagem , Humanos , Masculino , Pessoa de Meia-Idade , Nivolumabe/administração & dosagem , Estudos Retrospectivos , Fatores de Risco
6.
World J Gastroenterol ; 25(36): 5569-5577, 2019 Sep 28.
Artigo em Inglês | MEDLINE | ID: mdl-31576101

RESUMO

BACKGROUND: Intraductal papillary neoplasm of the bile duct (IPNB) is a type of tumor that presents in the intra- or extrahepatic bile ducts. Cystic-type intrahepatic IPNB often mimics simple liver cysts, making the diagnosis difficult. Because the growth of IPNB is slow, careful follow-up and timely therapeutic intervention is recommended. There are few reports with a follow-up period longer than a decade; thus, we report the case of a patient with an IPNB that grew for over 13 years. CASE SUMMARY: A 65-year-old man was diagnosed, 13 years prior with a cystic hepatic tumor with abnormal imaging findings. The targeted tumor biopsy results showed no malignancy. Biannual follow-up examinations were performed because of the potential for malignancy. The cystic lesions showed gradual enlargement over 11 years and a 4 mm papillary proliferation appeared on the cyst wall, which is compatible with IPNB. The tumor was observed for another 2 years because of the patient's wishes. The imaging findings showed enlargement to 8 mm and a new 9 mm papillary proliferation of the cystic tumor. Contrast-enhanced ultrasonography showed hyperenhancement during the arterial phase in both cyst walls, indicating intraductal tumor progression in both tumors. Thus, liver segment 8 subsegmentectomy was performed. The pathological findings indicated that the tumors contained mucin, and high-grade atypia was observed in the papillary lesions, showing IPNB. CONCLUSION: The development of IPNB should be monitored in patients with cystic lesions and ultrasonography are useful tool for the evaluation.


Assuntos
Neoplasias dos Ductos Biliares/diagnóstico , Ductos Biliares Intra-Hepáticos/patologia , Carcinoma Papilar/diagnóstico , Cistos/patologia , Idoso , Neoplasias dos Ductos Biliares/patologia , Neoplasias dos Ductos Biliares/cirurgia , Ductos Biliares Intra-Hepáticos/diagnóstico por imagem , Biópsia , Carcinoma Papilar/patologia , Carcinoma Papilar/cirurgia , Colangiografia , Cistos/cirurgia , Hepatectomia , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Tempo , Tomografia Computadorizada por Raios X , Resultado do Tratamento , Ultrassonografia
7.
JGH Open ; 3(4): 329-337, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31406927

RESUMO

BACKGROUND AND AIM: Several reports have demonstrated that skeletal muscle mass influences mortality in patients with hepatocellular carcinoma (HCC) receiving sorafenib treatment; however, there is still controversy with regard to whether skeletal muscle and adipose tissue are associated with the prognosis in HCC patients. We examined the relationship between body composition and prognosis in HCC patients. METHODS: We retrospectively analyzed 82 patients with unresectable HCC receiving sorafenib treatment. The skeletal muscle area and adipose tissue area were measured by computed tomography. Patients with low skeletal muscle index (male ≤36.2 cm2/m2, female ≤29.6 cm2/m2) and high visceral to subcutaneous adipose tissue area ratio (VSR) (male ≥ 1.33, female ≥ 0.93) were diagnosed as low skeletal muscle mass (LSMM) and high VSR, respectively. RESULTS: A total of 16 and 34 patients were classified as LSMM and high VSR, respectively. LSMM patients frequently experienced serious adverse events (SAEs) and thus had a shorter duration of sorafenib treatment than non-LSMM patients. High VSR was a significant factor for progression-free survival. LSMM patients less frequently received additional/subsequent therapies combined with sorafenib than non-LSMM patients. Multivariate Cox hazard analysis demonstrated that LSMM was a significant factor for the duration of sorafenib treatment. The treatment duration and receiving of additional/subsequent therapies were significantly associated with overall survival (OS) but not with LSMM or high VSR. CONCLUSION: LSMM was associated with the frequency of SAEs, treatment tolerability, and treatment duration. LSMM patients were less likely to receive additional/subsequent therapies than non-LSMM patients. Thus, LSMM could identify a subgroup of patients with poor OS.

8.
Intern Med ; 58(12): 1747-1752, 2019 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-30799364

RESUMO

The patient was a 76-year-old man who was treated with nivolumab due to recurrent gastric cancer. A blood examination revealed grade 3 alkaline phosphatase (ALP) elevation. A histopathological examination revealed marked portal infiltration, including eosinophils and CD4+ and CD8+ T lymphocytes, suggesting nivolumab-related cholangitis accompanied by the features of both an immune-related adverse event (irAE) and drug-induced liver injury (DILI) with allergic reaction. The patient's ALP level immediately decreased after the administration of prednisolone. Although nivolumab-related cholangitis, a rare irAE, has been reported to be refractory to steroid therapy, patients with features of irAE and allergic DILI might immediately respond to prednisolone.


Assuntos
Colangite/induzido quimicamente , Colangite/tratamento farmacológico , Nivolumabe/efeitos adversos , Prednisolona/uso terapêutico , Idoso , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Eosinófilos/metabolismo , Humanos , Fígado/patologia , Masculino , Nivolumabe/uso terapêutico , Prednisolona/administração & dosagem , Neoplasias Gástricas/tratamento farmacológico
9.
Cancer Sci ; 110(2): 674-685, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30548114

RESUMO

L-Type amino acid transporter 1 (LAT1) disulfide linked to CD98 heavy chain (hc) is highly expressed in most cancer cells, but weakly expressed in normal cells. In the present study, we developed novel anti-LAT1 mAbs and showed internalization activity, inhibitory effects of amino acid uptake and cell growth and antibody-dependent cellular cytotoxicity, as well as in vivo antitumor effects in athymic mice. Furthermore, we examined the reactivity of mAbs with LAT1 of Macaca fascicularis to evaluate possible side-effects of antihuman LAT1 mAbs in clinical trials. Antihuman LAT1 mAbs reacted with ACHN human and MK.P3 macaca kidney-derived cells, and this reactivity was significantly decreased by siRNAs against LAT1. Macaca LAT1 cDNA was cloned from MK.P3, and only two amino acid differences between human and macaca LAT1 were seen. RH7777 rat hepatoma and HEK293 human embryonic kidney cells expressing macaca LAT1 were established as stable transfectants, and antihuman LAT1 mAbs were equivalently reactive against transfectants expressing human or macaca LAT1. Dual (high and low) avidity modes were detected in transfectants expressing macaca LAT1, MK.P3, ACHN and HCT116 human colon cancer cells, and KA values were increased by anti-CD98hc mAb, suggesting anti-LAT1 mAbs detect an epitope on LAT1-CD98hc complexes on the cell surface. Based on these results, LAT1 may be a promising anticancer target and Macaca fascicularis can be used in preclinical studies with antihuman LAT1 mAbs.


Assuntos
Anticorpos Monoclonais/farmacologia , Transportador 1 de Aminoácidos Neutros Grandes/metabolismo , Células A549 , Aminoácidos/metabolismo , Animais , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Células HCT116 , Células HEK293 , Haplorrinos , Células HeLa , Humanos , Macaca fascicularis , Camundongos , Camundongos Nus , Ratos , Ratos Endogâmicos F344
10.
J Gastroenterol Hepatol ; 33(1): 283-290, 2018 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-28497593

RESUMO

BACKGROUND AND AIM: Some single-nucleotide polymorphisms (SNPs) are associated with the development of non-alcoholic fatty liver disease (NAFLD). As one of the genetic factors, PNPLA3 rs738409 (I148M) is important to associate with pathogenesis of NAFLD. Because other SNPs remain unclear in Japan, we performed a high-throughput sequencing that targeted more than 1000 genes to identify a novel genetic variant in Japanese patients with NAFLD. METHODS: The present study in 36 NAFLD patients and 27 healthy volunteers was performed. A high-throughput sequencer was used to detect the gene variations. Candidate genes were validated by TaqMan SNP genotyping assay in 53 NAFLD patients and 41 healthy volunteers. To investigate the function of candidate gene, we performed biochemical analyses in cultured hepatocytes and liver tissues. RESULTS: EXO1 rs1047840, PTPRD rs35929428, IFNAR2 rs2229207, CPOX rs1131857, IL23R rs1884444, IL10RA rs2228055, and FAM3B rs111988437 were identified as candidate genetic variants, and PTPRD rs35929428 was only extracted as a SNP predicting to cause protein dysfunction. In validation analysis, PTPRD rs35929428 associated with the development of NAFLD (P = 0.015, odds ratio = 5.00, 95% confidence interval: 1.33-18.70). In addition, PTPRD rs35929428 was associated with Fib-4 index and with hepatic fat droplets. Biochemical analyses indicated that PTPRD rs35929428 promoted dephosphorylation of tyrosine 705 signal transducer and activator of transcription 3 (Tyr 705) in hepatocytes. CONCLUSION: PTPRD rs35929428 was a novel SNP in patients with NAFLD. Through exacerbation of the dephosphorylation of signal transducer and activator of transcription 3 (Tyr 705) in hepatocytes, PTPRD rs35929428 might play a role in hepatic lipid accumulation and fibrosis, followed by the development of NAFLD.


Assuntos
Estudos de Associação Genética , Hepatopatia Gordurosa não Alcoólica/genética , Polimorfismo de Nucleotídeo Único/genética , Proteínas Tirosina Fosfatases Classe 2 Semelhantes a Receptores/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Povo Asiático/genética , Enzimas Reparadoras do DNA/genética , Exodesoxirribonucleases/genética , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Lipase/genética , Masculino , Proteínas de Membrana/genética , Pessoa de Meia-Idade , Receptor de Interferon alfa e beta/genética , Adulto Jovem
11.
J Biol Chem ; 287(11): 7896-906, 2012 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-22262832

RESUMO

We previously showed that depletion of the retinoblastoma protein (RB) induces down-regulation of the adhesion molecule E-cadherin and thereby triggers the epithelial-mesenchymal transition. To further characterize the effect of RB inactivation on the phenotype of cancer cells, we have now examined RB expression in human breast cancer cell lines and clinical specimens. We found that RB-inactive cells exhibit a mesenchymal-like morphology and are highly invasive. We also found that ZEB proteins, transcriptional repressors of the E-cadherin gene, are markedly up-regulated in these cells in a manner sensitive to the miR-200 family of microRNAs. Moreover, depletion of ZEB in RB-inactive cells suppressed cell invasiveness and proliferation and induced epithelial marker expression. These results implicate ZEB in induction of the epithelial-mesenchymal transition, as well as in maintenance of the mesenchymal phenotype in RB-inactive cells. We also developed a screening program for inhibitors of ZEB1 expression and thereby identified several cyclin-dependent kinase inhibitors that blocked both ZEB1 expression and RB phosphorylation. Together, our findings suggest that RB inactivation contributes to tumor progression not only through loss of cell cycle control but also through up-regulation of ZEB expression and induction of an invasive phenotype.


Assuntos
Biomarcadores Tumorais/biossíntese , Neoplasias da Mama/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas de Homeodomínio/biossíntese , Proteína do Retinoblastoma/biossíntese , Fatores de Transcrição/biossíntese , Biomarcadores Tumorais/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Caderinas/biossíntese , Caderinas/genética , Ciclo Celular/efeitos dos fármacos , Ciclo Celular/genética , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Feminino , Proteínas de Homeodomínio/genética , Humanos , MicroRNAs/biossíntese , MicroRNAs/genética , Invasividade Neoplásica , Inibidores de Proteínas Quinases/farmacologia , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Proteína do Retinoblastoma/genética , Fatores de Transcrição/genética , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco
12.
Int J Cancer ; 130(11): 2568-79, 2012 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-21717460

RESUMO

Triple-negative breast cancer [TNBC, which is negative for the estrogen receptor (ER), progesterone receptor, and human epidermal growth factor receptor 2] is a high-risk form of the disease without a specific therapy. DNA microarray and immunohistochemical analyses have shown that most TNBCs fall within the basal-like histological subset of breast cancers, which frequently exhibit inactivation of the retinoblastoma tumor suppressor (Rb) and upregulation of the cyclin-dependent kinase inhibitor p16(INK4a) (p16). However, downregulation of p16 expression has been observed in some basal-like breast cancer cell lines, suggesting that such cells can be divided into two groups according to Rb and p16 status. We now show that cells that are CD44(+) and CD24(-) , a phenotype associated with stem-like breast cancer cells, are more abundant in ER(-) /p16(-) breast cancer cell lines than in ER(-) /p16(+) lines. It was also found that p16 expression was downregulated in mammospheres from an ER-negative breast cancer cell line. Depletion of p16 by RNA interference in ER-negative breast cancer cells increased the percentage of CD44(+) /CD24(-) cells and increased the expression of mRNA of the ES-like genes Nanog, Oct4, and Sox2 through an Rb-independent pathway. Furthermore, such depletion of p16 reduced chemosensitivity. The loss of p16 expression may thus reduce the response of ER-negative breast cancer cells to chemotherapy by conferring cancer stem cell-like properties. Consistent with this conclusion, immunohistochemical analysis of the clinical samples suggests that low p16 expression in TNBC is associated with resistance to preoperative chemotherapy.


Assuntos
Neoplasias da Mama/tratamento farmacológico , Proteínas de Neoplasias/fisiologia , Adulto , Idoso , Apoptose/efeitos dos fármacos , Neoplasias da Mama/química , Neoplasias da Mama/patologia , Antígeno CD24/análise , Linhagem Celular Tumoral , Inibidor p16 de Quinase Dependente de Ciclina , Resistencia a Medicamentos Antineoplásicos , Feminino , Proteínas de Homeodomínio/genética , Humanos , Receptores de Hialuronatos/análise , Pessoa de Meia-Idade , Proteína Homeobox Nanog , Células-Tronco Neoplásicas , Fator 3 de Transcrição de Octâmero/genética , Paclitaxel/farmacologia , Fosforilação , Receptores de Estrogênio/análise , Proteína do Retinoblastoma/fisiologia , Fatores de Transcrição SOXB1/genética , Proteína Smad3/metabolismo
13.
Exp Dermatol ; 19(8): e136-41, 2010 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-20002172

RESUMO

Plexiform and/or dermal neurofibromas are nerve sheath tumors of the peripheral nervous system that are usually present in individuals with neurofibromatosis type 1 (NF1). Neurofibromas arise from Schwann cells with biallelic inactivation of NF1, the gene that encodes neurofibromin. This protein is responsible for regulation of the Ras-mediated pathway, which has been shown to play a crucial role in epithelial-to-mesenchymal transition (EMT). EMT is a biological process that occurs during embryogenesis and wound healing and is involved in pathological processes such as organ fibrosis and cancer metastasis. However, the relationship between neurofibromin and EMT has not been elucidated. We investigated whether the EMT-related signaling pathway was upregulated in NF1-associated neurofibromas and Schwann cells by assessing the expression levels of the EMT-related transcription factors Snail, Slug, Twist, ZEB1 and ZEB2. Immunohistochemical studies and quantitative reverse transcription polymerase chain reaction revealed an increase in the expression levels of EMT-related transcription factors in neurofibroma specimens and NF1-derived Schwann cells (sNF96.2). In addition, the silencing of NF1 by siRNA induced the expression of EMT-related transcription factors in normal human Schwann cells and in epithelial-like breast cancer cells. Our findings suggest that the loss of neurofibromin activated the EMT-related signaling pathway and that the excessive mesenchymal reaction may play a key role in the development of NF1-associated neurofibromas.


Assuntos
Diferenciação Celular/fisiologia , Células Epiteliais/patologia , Mesoderma/patologia , Neurofibromatose 1/metabolismo , Neurofibromatose 1/patologia , Neurofibromina 1/metabolismo , Transdução de Sinais/fisiologia , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Células Cultivadas , Células Epiteliais/metabolismo , Humanos , Mesoderma/metabolismo , Neoplasias de Bainha Neural/metabolismo , Neoplasias de Bainha Neural/patologia , Neurofibroma/metabolismo , Neurofibroma/patologia , Células de Schwann/metabolismo , Células de Schwann/patologia , Fatores de Transcrição/metabolismo
14.
Cancer Res ; 68(13): 5104-12, 2008 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-18593909

RESUMO

The retinoblastoma tumor suppressor protein (Rb) is mutated or expressed at very low levels in several tumor types, including retinoblastoma and osteosarcoma, as well as small cell lung, colon, prostate, bladder, and breast carcinomas. Loss or reduction of Rb expression is seen most commonly in high-grade breast adenocarcinomas, suggesting that a relationship may exist between loss of Rb function and a less-differentiated state, increased proliferation, and high metastatic potential. In this study, we found that knockdown of Rb by small interfering RNA in MCF7 breast cancer cells disrupts cell-cell adhesion and induces a mesenchymal-like phenotype. The epithelial-to-mesenchymal transition (EMT), a key event in embryonic morphogenesis, is implicated in the metastasis of primary tumors. Additionally, Rb is decreased during growth factor- and cytokine-induced EMT and overexpression of Rb inhibits the EMT in MCF10A human mammary epithelial cells. Ectopic expression and knockdown of Rb resulted in increased or reduced expression of E-cadherin, which is specifically involved in epithelial cell-cell adhesion. Other EMT-related transcriptional factors, including Slug and Zeb-1, are also induced by Rb depletion. Furthermore, we confirmed that Rb binds to an E-cadherin promoter sequence in association with the transcription factor activator protein-2alpha. Finally, in breast cancer specimens, we observed a concurrent down-regulation of Rb and E-cadherin expression in mesenchymal-like invasive cancers. These findings suggest that Rb inactivation contributes to tumor progression due to not only loss of cell proliferation control but also conversion to an invasive phenotype and that the inhibition of EMT is a novel tumor suppressor function of Rb.


Assuntos
Caderinas/genética , Células Epiteliais/fisiologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Mesoderma/fisiologia , RNA Interferente Pequeno/farmacologia , Proteína do Retinoblastoma/antagonistas & inibidores , Neoplasias da Mama/patologia , Caderinas/metabolismo , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Progressão da Doença , Desenvolvimento Embrionário/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Humanos , Mesoderma/efeitos dos fármacos , Mesoderma/metabolismo , Invasividade Neoplásica , Fenótipo , Proteína do Retinoblastoma/genética , Proteína do Retinoblastoma/fisiologia , Transfecção , Fator de Crescimento Transformador beta/farmacologia , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/farmacologia , Proteínas rho de Ligação ao GTP/metabolismo , Proteínas rho de Ligação ao GTP/fisiologia
15.
Nat Struct Mol Biol ; 15(3): 228-36, 2008 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-18278057

RESUMO

Eukaryotic 20S proteasomes are composed of two alpha-rings and two beta-rings, which form an alphabetabetaalpha stacked structure. Here we describe a proteasome-specific chaperone complex, designated Dmp1-Dmp2, in budding yeast. Dmp1-Dmp2 directly bound to the alpha5 subunit to facilitate alpha-ring formation. In Deltadmp1 cells, alpha-rings lacking alpha4 and decreased formation of 20S proteasomes were observed. Dmp1-Dmp2 interacted with proteasome precursors early during proteasome assembly and dissociated from the precursors before the formation of half-proteasomes. Notably, the crystallographic structures of Dmp1 and Dmp2 closely resemble that of PAC3-a mammalian proteasome-assembling chaperone; nonetheless, neither Dmp1 nor Dmp2 showed obvious sequence similarity to PAC3. The structure of the Dmp1-Dmp2-alpha5 complex reveals how this chaperone functions in proteasome assembly and why it dissociates from proteasome precursors before the beta-rings are assembled.


Assuntos
Chaperonas Moleculares/química , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/enzimologia , Cristalografia por Raios X , Precursores Enzimáticos/metabolismo , Chaperonas Moleculares/metabolismo , Complexos Multienzimáticos/química , Complexos Multienzimáticos/isolamento & purificação , Complexos Multienzimáticos/metabolismo , Proteínas Mutantes/química , Proteínas Mutantes/isolamento & purificação , Proteínas Mutantes/metabolismo , Mutação/genética , Complexo de Endopeptidases do Proteassoma/química , Ligação Proteica , Estrutura Secundária de Proteína , Saccharomyces cerevisiae/citologia , Proteínas de Saccharomyces cerevisiae/metabolismo , Homologia Estrutural de Proteína
16.
Science ; 316(5829): 1349-53, 2007 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-17540904

RESUMO

Proteasomes are responsible for generating peptides presented by the class I major histocompatibility complex (MHC) molecules of the immune system. Here, we report the identification of a previously unrecognized catalytic subunit called beta5t. beta5t is expressed exclusively in cortical thymic epithelial cells, which are responsible for the positive selection of developing thymocytes. Although the chymotrypsin-like activity of proteasomes is considered to be important for the production of peptides with high affinities for MHC class I clefts, incorporation of beta5t into proteasomes in place of beta5 or beta5i selectively reduces this activity. We also found that beta5t-deficient mice displayed defective development of CD8(+) T cells in the thymus. Our results suggest a key role for beta5t in generating the MHC class I-restricted CD8(+) T cell repertoire during thymic selection.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Timo/citologia , Timo/enzimologia , Sequência de Aminoácidos , Animais , Autoantígenos/imunologia , Autoantígenos/metabolismo , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/citologia , Domínio Catalítico , Células Epiteliais/enzimologia , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Linfopoese , Dados de Sequência Molecular , Peptídeos/química , Peptídeos/metabolismo , Complexo de Endopeptidases do Proteassoma/química , Complexo de Endopeptidases do Proteassoma/genética , Baço/citologia , Baço/imunologia , Subpopulações de Linfócitos T/citologia , Subpopulações de Linfócitos T/imunologia , Timo/imunologia
17.
Mol Cell ; 24(6): 977-84, 2006 Dec 28.
Artigo em Inglês | MEDLINE | ID: mdl-17189198

RESUMO

The 20S proteasome is a catalytic core of the 26S proteasome, a central enzyme in the degradation of ubiquitin-conjugated proteins. It is composed of 14 distinct gene products that form four stacked rings of seven subunits each, alpha(1-7)beta(1-7)beta(1-7)alpha(1-7). It is reported that the biogenesis of mammalian 20S proteasomes is assisted by proteasome-specific chaperones, named PAC1, PAC2, and hUmp1, but the details are still unknown. Here, we report the identification of a chaperone, designated PAC3, as a component of alpha rings. Although it can intrinsically bind directly to both alpha and beta subunits, PAC3 dissociates before the formation of half-proteasomes, a process coupled with the recruitment of beta subunits and hUmp1. Knockdown of PAC3 impaired alpha ring formation. Further, PAC1/2/3 triple knockdown resulted in the accumulation of disorganized half-proteasomes that are incompetent for dimerization. Our results describe a cooperative system of multiple chaperones involved in the correct assembly of mammalian 20S proteasomes.


Assuntos
Chaperonas Moleculares/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Linhagem Celular , Humanos , Modelos Biológicos , Chaperonas Moleculares/genética , Complexo de Endopeptidases do Proteassoma/biossíntese , RNA Interferente Pequeno
18.
J Biochem ; 135(2): 185-91, 2004 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-15047720

RESUMO

Escherichia coli spr (suppressor of prc) mutants and nlpI mutants show thermosensitive growth. The thermosensitivity of the spr mutants was suppressed by the nlpI mutations. Expression of the fusion genes encoding hexa-histidine-tagged NlpI (NlpI-His) and purification of the tagged NlpI showed that NlpI-His bound with Prc protease and IbpB chaperone. NlpI-His with the amino acid substitution of G103D did not bind with either of these proteins, while NlpI-His variants (NlpI-284-His, NlpI-Q283-His, and NlpI-G282-His) lacking 10 to 12 residues from the carboxy terminus bound with both proteins. The tagged NlpI lacking 11 amino acid residues from the carboxy terminus was processed by Prc, but that lacking 12 residues was not. The thermosensitivity of the nlpI mutant was corrected by the production of the former NlpI variant, but not by production of the latter. Expression of the truncated NlpI that lacked 10 or 11 residues from the carboxy terminus corrected the thermosensitivity of the prc nlpI double mutant, while expression of the full-length NlpI did not. Thus, it was suggested that NlpI was activated by Prc protease processing.


Assuntos
Endopeptidases/metabolismo , Proteínas de Escherichia coli/metabolismo , Escherichia coli/metabolismo , Lipoproteínas/metabolismo , Sequência de Aminoácidos , Sequência de Bases , Temperatura Alta , Lipoproteínas/química , Dados de Sequência Molecular , Mutação , Plasmídeos/genética , Fatores de Tempo
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