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1.
Carcinogenesis ; 45(3): 119-130, 2024 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-38123365

RESUMO

The role of the ferroptosis-related gene glutathione peroxidase 4 (GPX4) in oncology has been extensively investigated. However, the clinical implications of GPX4 in patients with intrahepatic cholangiocarcinoma (ICC) remain unknown. This study aimed to evaluate the prognostic impact of GPX4 and its underlying molecular mechanisms in patients with ICC. Fifty-seven patients who underwent surgical resection for ICC between 2010 and 2017 were retrospectively analyzed. Based on the immunohistochemistry, patients were divided into GPX4 high (n = 15) and low (n = 42) groups, and clinical outcomes were assessed. Furthermore, the roles of GPX4 in cell proliferation, migration and gene expression were analyzed in ICC cell lines in vitro and in vivo. The results from clinical study showed that GPX4 high group showed significant associations with high SUVmax on 18F-fluorodeoxyglucose-positron emission tomography (≥8.0, P = 0.017), multiple tumors (P = 0.004), and showed glucose transporter 1 (GLUT1) high expression with a trend toward significance (P = 0.053). Overall and recurrence-free survival in the GPX4 high expression group were significantly worse than those in the GPX4 low expression group (P = 0.038 and P < 0.001, respectively). In the experimental study, inhibition of GPX4 attenuated cell proliferation and migration in ICC cell lines. Inhibition of GPX4 also decreased the expression of glucose metabolism-related genes, such as GLUT1 or HIF1α. Mechanistically, these molecular changes are regulated in Akt-mechanistic targets of rapamycin axis. In conclusion, this study suggested the pivotal value of GPX4 serving as a prognostic marker for patients with ICC. Furthermore, GPX4 can mediate glucose metabolism of ICC.


Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Ferroptose , Humanos , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/genética , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ferroptose/genética , Transportador de Glucose Tipo 1/genética , Estudos Retrospectivos , Colangiocarcinoma/genética , Colangiocarcinoma/cirurgia , Colangiocarcinoma/metabolismo , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Ductos Biliares Intra-Hepáticos/patologia , Neoplasias dos Ductos Biliares/patologia , Glucose
2.
Hepatology ; 77(2): 443-455, 2023 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-35603471

RESUMO

BACKGROUND AND AIMS: The mechanism underlying liver regeneration following partial hepatectomy (PH) is not fully elucidated. We aimed to characterize collagen gene expressing hepatic cells following PH and examine their contribution to liver regeneration. APPROACH AND RESULTS: Col-GFP mice, which express GFP under the control of the collagen gene promoter, were used to detect collagen gene expressing cells following PH. The GFP-expressing cells were analyzed via single-cell RNA sequencing (scRNA-seq). Additionally, Col-ER Cre/RFP and Col-ER Cre/DTA mice were utilized to examine the cell fates and functional roles of collagen gene expressing cells in liver regeneration, respectively. The number of collagen gene expressing cells was found to be increased on day 3 and subsequently decreased on day 7 following PH. ScRNA-seq analysis of sorted collagen gene expressing cells showed that the regenerating liver was characterized by three distinct hepatic stellate cell (HSC) clusters, including one representing classic myofibroblasts. The other HSC clusters included an intermediately activated HSC cluster and a proliferating HSC cluster. Of these, the latter cluster was absent in the CCl 4 -induced liver fibrosis model. Cell fate tracing analysis using Col-ER Cre/RFP mice demonstrated that the collagen gene expressing cells escaped death during regeneration and remained in an inactivated state in the liver. Further, depletion of these cells using Col-ER Cre/DTA mice resulted in impaired liver regeneration. CONCLUSIONS: Heterogeneous HSC clusters, one of which was a unique proliferating cluster, were found to appear in the liver following PH. Collagen gene expressing cells, including HSCs, were found to promote liver regeneration.


Assuntos
Hepatectomia , Hepatócitos , Camundongos , Animais , Hepatócitos/metabolismo , Fígado/metabolismo , Cirrose Hepática/patologia , Células Estreladas do Fígado/metabolismo , Colágeno/metabolismo
3.
Hepatol Res ; 54(9): 817-826, 2024 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-38430513

RESUMO

BACKGROUND AND AIM: Autotaxin (ATX) is an extracellular lysophospholipase D that catalyzes the hydrolysis of lysophosphatidylcholine into lysophosphatidic acid (LPA). Recent accumulating evidence indicates the biological roles of ATX in malignant tumors. However, the expression and clinical implications of ATX in human cholangiocarcinoma (CCA) remain elusive. METHODS: In this study, the expression of ATX in 97 human CCA tissues was evaluated by immunohistochemistry. Serum ATX levels were determined in CCA patients (n = 26) and healthy subjects (n = 8). Autotaxin expression in cell types within the tumor microenvironment was characterized by immunofluorescence staining. RESULTS: High ATX expression in CCA tissue was significantly associated with a higher frequency of lymph node metastasis (p = 0.050). High ATX expression was correlated with shorter overall survival (p = 0.032) and recurrence-free survival (RFS) (p = 0.001) than low ATX expression. In multivariate Cox analysis, high ATX expression (p = 0.019) was an independent factor for shorter RFS. Compared with low ATX expression, high ATX expression was significantly associated with higher Ki-67-positive cell counts (p < 0.001). Serum ATX levels were significantly higher in male CCA patients than in healthy male subjects (p = 0.030). In the tumor microenvironment of CCA, ATX protein was predominantly expressed in tumor cells, cancer-associated fibroblasts, plasma cells, and biliary epithelial cells. CONCLUSIONS: Our study highlights the clinical evidence and independent prognostic value of ATX in human CCA.

4.
Proc Natl Acad Sci U S A ; 118(29)2021 07 20.
Artigo em Inglês | MEDLINE | ID: mdl-34253615

RESUMO

We investigated the role of mesothelin (Msln) and thymocyte differentiation antigen 1 (Thy1) in the activation of fibroblasts across multiple organs and demonstrated that Msln-/- mice are protected from cholestatic fibrosis caused by Mdr2 (multidrug resistance gene 2) deficiency, bleomycin-induced lung fibrosis, and UUO (unilateral urinary obstruction)-induced kidney fibrosis. On the contrary, Thy1-/- mice are more susceptible to fibrosis, suggesting that a Msln-Thy1 signaling complex is critical for tissue fibroblast activation. A similar mechanism was observed in human activated portal fibroblasts (aPFs). Targeting of human MSLN+ aPFs with two anti-MSLN immunotoxins killed fibroblasts engineered to express human mesothelin and reduced collagen deposition in livers of bile duct ligation (BDL)-injured mice. We provide evidence that antimesothelin-based therapy may be a strategy for treatment of parenchymal organ fibrosis.


Assuntos
Colestase/tratamento farmacológico , Fibroblastos/imunologia , Imunoterapia , Cirrose Hepática/tratamento farmacológico , Animais , Colestase/genética , Colestase/imunologia , Colágeno/imunologia , Fibroblastos/efeitos dos fármacos , Humanos , Imunotoxinas/administração & dosagem , Cirrose Hepática/genética , Cirrose Hepática/imunologia , Mesotelina/genética , Mesotelina/imunologia , Camundongos , Antígenos Thy-1/genética , Antígenos Thy-1/imunologia
5.
Hepatol Res ; 53(2): 145-159, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36149410

RESUMO

AIM: Sinusoidal obstruction syndrome (SOS) induced by oxaliplatin-including chemotherapies (OXCx) is associated with impaired hepatic reserve and higher morbidity after hepatic resection. However, in the absence of an appropriate animal experimental model, little is known about its pathophysiology. This study aimed to establish a clinically relevant reproducible model of FOLFOX-induced SOS and to compare the clinical/histopathological features between the clinical and animal SOS settings. METHODS: We performed clinical/pathological analyses of colorectal liver metastasis (CRLM) patients who underwent hepatectomy with/without preoperative treatment of FOLFOX (n = 22/18). Male micro-minipigs were treated with 50% of the standard human dosage of the FOLFOX regimen. RESULTS: In contrast to the monocrotaline-induced SOS model in rats, hepatomegaly, ascites, congestion, and coagulative necrosis of hepatocytes were absent in patients with CRLM with OXCx pretreatment and OXCx-treated micro-minipigs. In parallel to CRLM cases with OXCx pretreatment, OXCx-challenged micro-minipigs exhibited deteriorated indocyanine green clearance, morphological alteration of liver sinusoidal endothelial cells, and upregulated matrix metalloproteinase-9. Using our novel porcine SOS model, we identified the hepatoprotective influence of recombinant human soluble thrombomodulin in OXCx-SOS. CONCLUSIONS: With distinct differences between monocrotaline-induced rat SOS and human/pig OXCx-SOS, our pig OXCx-SOS model serves as a preclinical platform for future investigations to dissect the pathophysiology of OXCx-SOS and seek preventive strategies.

6.
Cancer Sci ; 113(1): 41-52, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34716967

RESUMO

Due to the considerable success of cancer immunotherapy for leukemia, the tumor immune environment has become a focus of intense research; however, there are few reports on the dynamics of the tumor immune environment in leukemia. Here, we analyzed the tumor immune environment in pediatric B cell precursor acute lymphoblastic leukemia by analyzing serial bone marrow samples from nine patients with primary and recurrent disease by mass cytometry using 39 immunophenotype markers, and transcriptome analysis. High-dimensional single-cell mass cytometry analysis elucidated a dynamic shift of T cells from naïve to effector subsets, and clarified that, during relapse, the tumor immune environment comprised a T helper 1-polarized immune profile, together with an increased number of effector regulatory T cells. These results were confirmed in a validation cohort using conventional flow cytometry. Furthermore, RNA transcriptome analysis identified the upregulation of immune-related pathways in B cell precursor acute lymphoblastic leukemia cells during relapse, suggesting interaction with the surrounding environment. In conclusion, a tumor immune environment characterized by a T helper 1-polarized immune profile, with an increased number of effector regulatory T cells, could contribute to the pathophysiology of recurrent B cell precursor acute lymphoblastic leukemia. This information could contribute to the development of effective immunotherapeutic approaches against B cell precursor acute lymphoblastic leukemia relapse.


Assuntos
Biomarcadores Tumorais/genética , Medula Óssea/imunologia , Perfilação da Expressão Gênica/métodos , Recidiva Local de Neoplasia/genética , Leucemia-Linfoma Linfoblástico de Células Precursoras B/genética , Adolescente , Medula Óssea/química , Criança , Pré-Escolar , Feminino , Citometria de Fluxo , Regulação Neoplásica da Expressão Gênica , Humanos , Lactente , Masculino , Recidiva Local de Neoplasia/imunologia , Leucemia-Linfoma Linfoblástico de Células Precursoras B/imunologia , Análise de Sequência de RNA , Análise de Célula Única , Microambiente Tumoral , Regulação para Cima , Adulto Jovem
7.
Liver Transpl ; 28(4): 647-658, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-34655506

RESUMO

Programmed death 1 (PD1)/its ligand PD-L1 concomitant with T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3)/its ligand galectin 9 (Gal-9) and the forkhead box P3 (FOXP3) might be involved in tolerance after liver transplantation (LT). Liver biopsies from 38 tolerant, 19 nontolerant (including 16 samples that triggered reintroduction of immunosuppression [IS] and 19 samples after IS reintroduction), and 38 control LT patients were studied. The expressions of PD1, PD-L1, Gal-9, and FOXP3 were determined by immunohistochemical and immunofluorescence (IF) staining. The success period of IS withdrawal was calculated using Kaplan-Meier curve analysis. Tolerant and control patients exhibited higher PD-L1, Gal-9, and FOXP3 levels than nontolerant patients at the moment of triggering IS reintroduction. High expressions of PD-L1 and Gal-9 were associated with prolonged success of tolerance (83.3% versus 36.7% [P < 0.01] and 73.1% versus 42.9% [P = 0.03]). A strong correlation between PD-L1 and Gal-9 expression levels was detected (Spearman r = 0.73; P ≤ 0.001), and IF demonstrated colocalization of PD-L1 and Gal-9 in the cytoplasm of hepatocytes. In conclusion, the present study demonstrated that increased expressions of PD-L1 and Gal-9 were associated with sustained tolerance after IS withdrawal in pediatric liver transplantation.


Assuntos
Antígeno B7-H1 , Transplante de Fígado , Antígeno B7-H1/análise , Antígeno B7-H1/metabolismo , Criança , Fatores de Transcrição Forkhead/análise , Galectinas/metabolismo , Humanos , Terapia de Imunossupressão/efeitos adversos , Ligantes , Transplante de Fígado/efeitos adversos , Transplantados
8.
Liver Transpl ; 27(3): 403-415, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-32780942

RESUMO

Mac-2 binding protein glycosylation isomer (M2BPGi) is a novel liver fibrosis biomarker, but there are few studies on M2BPGi in liver transplantation (LT) recipients. This study aimed to evaluate the utility of M2BPGi measurement in LT recipients. We collected the clinicopathological data of 233 patients who underwent a liver biopsy at Kyoto University Hospital after LT between August 2015 and June 2019. The median values of M2BPGi in patients with METAVIR fibrosis stages F0, F1, F2, and ≥F3 were 0.61, 0.76, 1.16, and 1.47, respectively, whereas those in patients with METAVIR necroinflammatory indexes A0, A1, and ≥A2 were 0.53, 1.145, and 2.24, respectively. Spearman rank correlation test suggested that the necroinflammatory index had a stronger correlation to the M2BPGi value than the fibrosis stage. The area under the receiver operating characteristic curve of M2BPGi to predict ≥A1 was 0.75, which was significantly higher than that of any other liver fibrosis and inflammation marker. Patients with a rejection activity index (RAI) of ≥3 had a higher M2BPGi value than those with RAI ≤ 2 (P = 0.001). Patients with hepatitis C virus viremia had a higher M2BPGi value than sustained virological responders or those with other etiologies. In conclusion, the present study demonstrated that M2BPGi values are more strongly influenced by necroinflammatory activity and revealed M2BPGi, which has been thought to be a so-called fibrosis marker, as a disease activity marker in transplant recipients. M2BPGi measurement may be useful to detect early stage liver inflammation that cannot be detected by routine blood examination of LT recipients.


Assuntos
Transplante de Fígado , Glicosilação , Humanos , Cirrose Hepática/diagnóstico , Cirrose Hepática/cirurgia , Transplante de Fígado/efeitos adversos , Glicoproteínas de Membrana/metabolismo , Curva ROC
9.
J Gastroenterol Hepatol ; 36(9): 2493-2500, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-33448457

RESUMO

BACKGROUND AND AIM: Studies on the resolution of liver fibrosis are becoming more important in this era of etiologic eradication. In contrast to the extensive research on the recovery of liver fibrosis induced by hepatotoxic injuries, regression of cholestatic liver fibrosis has been insufficiently examined owing to the limited availability of animal models. METHODS: We examined our novel recanalization mice model of biliary obstruction, involving anastomosis between the gallbladder and jejunum (G-J anastomosis) by invagination. Transgenic mice expressing green fluorescent protein (GFP) under the collagen 1(α)1 promoter underwent G-J anastomosis 14 days after bile duct ligation (BDL) and were sacrificed 14 days later. RESULTS: Transaminase and total bilirubin levels decreased to almost normal values on day 14 after G-J anastomosis. G-J anastomosis resulted in dramatic reversal of liver fibrosis induced by BDL. Activated portal fibroblasts (PFs) double-positive for GFP and Thy-1 on immunofluorescence in the liver of BDL-injured mice became less noticeable following G-J anastomosis. Messenger RNA expression of markers for activated PFs in the liver was downregulated after anastomosis. Matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) were induced by BDL. After anastomosis, expressions of MMP-3, 8 as well as hepatocyte growth factor were further upregulated, whereas those of TIMP-1 and TIMP-3 were markedly downregulated. CONCLUSIONS: Our established G-J anastomosis model is associated with fibrosis resolution and reduced PF activation through reopening of bile duct obstruction and will be valuable for studying the recovery process of cholestatic liver fibrosis.


Assuntos
Colestase , Vesícula Biliar , Anastomose Cirúrgica , Animais , Ductos Biliares/cirurgia , Colestase/etiologia , Colestase/patologia , Modelos Animais de Doenças , Ligadura , Fígado/patologia , Cirrose Hepática/etiologia , Cirrose Hepática/patologia , Camundongos , Camundongos Transgênicos
10.
Br J Cancer ; 122(2): 245-257, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31819188

RESUMO

BACKGROUND: The fatty acid (FA) composition of phosphatidylinositols (PIs) is tightly regulated in mammalian tissue since its disruption impairs normal cellular functions. We previously found its significant alteration in breast cancer by using matrix-assisted laser desorption and ionisation imaging mass spectrometry (MALDI-IMS). METHODS: We visualised the histological distribution of PIs containing different FAs in 65 primary breast cancer tissues using MALDI-IMS and investigated its association with clinicopathological features and gene expression profiles. RESULTS: Normal ductal cells (n = 7) predominantly accumulated a PI containing polyunsaturated FA (PI-PUFA), PI(18:0/20:4). PI(18:0/20:4) was replaced by PIs containing monounsaturated FA (PIs-MUFA) in all non-invasive cancer cells (n = 12). While 54% of invasive cancer cells (n = 27) also accumulated PIs-MUFA, 46% of invasive cancer cells (n = 23) accumulated the PIs-PUFA, PI(18:0/20:3) and PI(18:0/20:4). The accumulation of PI(18:0/20:3) was associated with higher incidence of lymph node metastasis and activation of the PD-1-related immune checkpoint pathway. Fatty acid-binding protein 7 was identified as a putative molecule controlling PI composition. CONCLUSIONS: MALDI-IMS identified PI composition associated with invasion and nodal metastasis of breast cancer. The accumulation of PI(18:0/20:3) could affect the PD-1-related immune checkpoint pathway, although its precise mechanism should be further validated.


Assuntos
Neoplasias da Mama/genética , Ácidos Graxos/metabolismo , Fosfatidilinositóis/metabolismo , Transcriptoma/genética , Idoso , Animais , Neoplasias da Mama/diagnóstico por imagem , Neoplasias da Mama/patologia , Ácidos Graxos/genética , Feminino , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Espectrometria de Massas , Pessoa de Meia-Idade , Fosfatidilinositóis/genética , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos
11.
Am J Transplant ; 20(3): 808-816, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31566887

RESUMO

This study aimed to evaluate postoperative long-term liver restoration and splenic enlargement and their clinical significance in living donor liver transplantation. One hundred and sixteen donors who had donated livers more than 5 years previously accepted the invitation to participate in this study. The liver restoration rate and the splenic enlargement rate were calculated as the rate with respect to the original volume. The mean liver restoration rate was 0.99 ± 0.12 and older age was associated with a higher incidence for liver restoration rate <0.95 (P = .005), whereas type of donor operation was not. The donors with liver restoration rate <0.95 showed lower serum albumin levels than those with liver restoration rate ≥0.95. The mean splenic enlargement rate was 1.10 ± 0.16. Right lobe donors demonstrated higher splenic enlargement rate (1.14 ± 0.18) than left lobe/lateral segment donors (1.06 ± 0.13). In the donors with splenic enlargement rate ≥1.10, platelet count was not fully restored to the preoperative level. In conclusion, older age increases the risk for incomplete postoperative liver restoration, which may be associated with a decrease in albumin more than 5 years after donation. Right lobe donation poses a risk of splenic enlargement, which is associated with incomplete restoration of platelet count.


Assuntos
Transplante de Fígado , Doadores Vivos , Idoso , Hepatectomia , Humanos , Fígado/cirurgia , Estudos Prospectivos , Baço
12.
Proc Natl Acad Sci U S A ; 114(38): 10178-10183, 2017 09 19.
Artigo em Inglês | MEDLINE | ID: mdl-28878025

RESUMO

The commensal microbiota within the gastrointestinal tract is essential in maintaining homeostasis. Indeed, dysregulation in the repertoire of microbiota can result in the development of intestinal immune-inflammatory diseases. Further, this immune regulation by gut microbiota is important systemically, impacting health and disease of organ systems beyond the local environment of the gut. What has not been explored is how distant organs might in turn shape the microbiota via microbe-targeted molecules. Here, we provide evidence that surfactant protein D (SP-D) synthesized in the gallbladder and delivered into intestinal lumen binds selectively to species of gut commensal bacteria. SP-D-deficient mice manifest intestinal dysbiosis and show a susceptibility to dextran sulfate sodium-induced colitis. Further, fecal transfer from SP-D-deficient mice to wild-type, germ-free mice conveyed colitis susceptibility. Interestingly, colitis caused a notable increase in Sftpd gene expression in the gallbladder, but not in the lung, via the activity of glucocorticoids produced in the liver. These findings describe a unique mechanism of interorgan regulation of intestinal immune homeostasis by SP-D with potential clinical implications such as cholecystectomy.


Assuntos
Colite/metabolismo , Vesícula Biliar/metabolismo , Microbioma Gastrointestinal , Proteína D Associada a Surfactante Pulmonar/metabolismo , Animais , Colite/microbiologia , Fatores de Transcrição Forkhead/metabolismo , Glucocorticoides/biossíntese , Homeostase , Mucosa Intestinal/imunologia , Fígado/metabolismo , Camundongos Endogâmicos C57BL , Simbiose , Linfócitos T Reguladores/metabolismo
13.
J Hepatol ; 71(3): 573-585, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31071368

RESUMO

BACKGROUND & AIMS: Chronic liver injury often results in the activation of hepatic myofibroblasts and the development of liver fibrosis. Hepatic myofibroblasts may originate from 3 major sources: hepatic stellate cells (HSCs), portal fibroblasts (PFs), and fibrocytes, with varying contributions depending on the etiology of liver injury. Here, we assessed the composition of hepatic myofibroblasts in multidrug resistance gene 2 knockout (Mdr2-/-) mice, a genetic model that resembles primary sclerosing cholangitis in patients. METHODS: Mdr2-/- mice expressing a collagen-GFP reporter were analyzed at different ages. Hepatic non-parenchymal cells isolated from collagen-GFP Mdr2-/- mice were sorted based on collagen-GFP and vitamin A. An NADPH oxidase (NOX) 1/4 inhibitor was administrated to Mdr2-/- mice aged 12-16 weeks old to assess the therapeutic approach of targeting oxidative stress in cholestatic injury. RESULTS: Thy1+ activated PFs accounted for 26%, 51%, and 54% of collagen-GFP+ myofibroblasts in Mdr2-/- mice at 4, 8, and 16 weeks of age, respectively. The remaining collagen-GFP+ myofibroblasts were composed of activated HSCs, suggesting that PFs and HSCs are both activated in Mdr2-/- mice. Bone-marrow-derived fibrocytes minimally contributed to liver fibrosis in Mdr2-/- mice. The development of cholestatic liver fibrosis in Mdr2-/- mice was associated with early recruitment of Gr1+ myeloid cells and upregulation of pro-inflammatory cytokines (4 weeks). Administration of a NOX inhibitor to 12-week-old Mdr2-/- mice suppressed the activation of myofibroblasts and attenuated the development of cholestatic fibrosis. CONCLUSIONS: Activated PFs and activated HSCs contribute to cholestatic fibrosis in Mdr2-/- mice, and serve as targets for antifibrotic therapy. LAY SUMMARY: Activated portal fibroblasts and hepatic stellate cells, but not fibrocytes, contributed to the production of the fibrous scar in livers of Mdr2-/- mice, and these cells can serve as targets for antifibrotic therapy in cholestatic injury. Therapeutic inhibition of the enzyme NADPH oxidase (NOX) in Mdr2-/- mice reversed cholestatic fibrosis, suggesting that targeting NOXs may be an effective strategy for the treatment of cholestatic fibrosis.


Assuntos
Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Fibroblastos/metabolismo , Células Estreladas do Fígado/metabolismo , Cirrose Hepática Biliar/metabolismo , Veia Porta/patologia , Animais , Células Cultivadas , Modelos Animais de Doenças , Técnicas de Inativação de Genes , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática Biliar/tratamento farmacológico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/metabolismo , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirazolonas , Piridinas/farmacologia , Piridinas/uso terapêutico , Piridonas , Membro 4 da Subfamília B de Transportadores de Cassetes de Ligação de ATP
14.
Biochem Biophys Res Commun ; 503(2): 544-549, 2018 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-29800570

RESUMO

Osteoclasts play a crucial role in osteolytic bone diseases, such as osteoporosis, rheumatoid arthritis, periodontitis, Paget's disease of bone and bone metastatic tumors. Therefore, controlling osteoclast differentiation and function has been considered a promising therapeutic strategy. Here, we show that necrostatin (Nec)-7, an inhibitor of programmed necrosis, strongly suppressed receptor activator of nuclear factor (NF)-κB ligand (RANKL)-induced osteoclastogenesis and bone resorption, without compromising macrophage colony-stimulating factor (M-CSF)-supported survival and growth of osteoclast precursor cells. Accordingly, Nec-7 significantly decreased the levels of RANKL-induced osteoclastogenic marker genes, such as cathepsin K. Mechanistically, Nec-7 neither affected MAPK nor NF-κB activation; however, it strongly inhibited the RANKL receptor (RANK) to nuclear factor of activated T cells c1 (NFATc1) signaling. Lentiviral expression of RANK in bone marrow-derived macrophages significantly restored osteoclastogenesis and NFATc1 amplification in Nec-7-treated cells. In this study, we revealed that Nec-7-sensitive pathways are crucially involved in osteoclast formation and function. Investigation of the molecular mechanism(s) through which Nec-7 inhibits RANK-NFATc1 signaling axis may lead to the development of new therapeutic strategies for bone disease.


Assuntos
Diferenciação Celular/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Fatores de Transcrição NFATC/metabolismo , Osteoclastos/efeitos dos fármacos , Receptor Ativador de Fator Nuclear kappa-B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiazóis/farmacologia , Animais , Reabsorção Óssea/tratamento farmacológico , Reabsorção Óssea/metabolismo , Células Cultivadas , Feminino , Macrófagos/citologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , Osteoclastos/citologia , Osteoclastos/metabolismo
15.
Liver Transpl ; 24(5): 645-654, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29524333

RESUMO

Ischemia/reperfusion injury (IRI) is one of the main causes of liver dysfunction after liver surgery. Involvement of endoplasmic reticulum (ER) stress in various diseases has been demonstrated, and CAAT/enhancer binding protein-homologous protein (CHOP) is a transcriptional regulator that is induced by ER stress. It is also a key regulator of ER stress-mediated apoptosis. The aim of this study was to investigate the role of CHOP in liver IRI. Wild type (WT) and CAAT/enhancer binding protein-homologous protein knockout (CHOP-/-) mice were subjected to 70% liver warm ischemia/reperfusion for 60 minutes. At different times after reperfusion, liver tissues and blood samples were collected for evaluation. Induction of ER stress including CHOP expression was ascertained. Liver damage was evaluated based on serum liver enzymes, liver histology, and neutrophil infiltration. Hepatocyte death including apoptosis was assessed. Liver warm IRI induced ER stress in both WT and CHOP-/- mice. In addition, CHOP expression was up-regulated in WT mice. At 6 hours after reperfusion, liver damage was attenuated in CHOP-/- mice. On the basis of terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining, apoptotic and necrotic cells were significantly reduced in CHOP-/- mice. CHOP deficiency also reduced the cleavage of caspase 3 and expression of the proapoptotic protein B cell lymphoma 2-associated X protein. Liver IRI induces CHOP expression, and CHOP deficiency attenuates liver IRI by inhibiting apoptosis. Elucidation of the function of CHOP in liver IRI may contribute to further investigation for a therapy against liver IRI associated with the ER stress pathway. Liver Transplantation 24 645-654 2018 AASLD.


Assuntos
Hepatócitos/metabolismo , Hepatopatias/prevenção & controle , Fígado/metabolismo , Traumatismo por Reperfusão/prevenção & controle , Fator de Transcrição CHOP/deficiência , Animais , Apoptose , Caspase 3/metabolismo , Modelos Animais de Doenças , Estresse do Retículo Endoplasmático , Predisposição Genética para Doença , Hepatócitos/patologia , Fígado/patologia , Hepatopatias/genética , Hepatopatias/metabolismo , Hepatopatias/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Necrose , Infiltração de Neutrófilos , Fenótipo , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/patologia , Fator de Transcrição CHOP/genética , Proteína X Associada a bcl-2/metabolismo
16.
J Transl Med ; 16(1): 95, 2018 04 11.
Artigo em Inglês | MEDLINE | ID: mdl-29642912

RESUMO

BACKGROUND: Surgical resection remains the mainstay of curative treatment for intrahepatic cholangiocarcinoma (ICC). Prognosis after surgery is unsatisfactory despite improvements in treatment and post-operative clinical management. Despite developments in the molecular profiling of ICC, the preoperative prediction of prognosis remains a challenge. This study aimed to identify clinical prognostic indicators by investigating the molecular profiles of ICC and evaluating the preoperative imaging data of 18F-fluorodeoxyglucose positron emission tomography (18F-FDG-PET). METHODS: A retrospective analysis was performed on 50 consecutive patients with ICC who underwent curative hepatectomy after 18F-FDG-PET examination. To evaluate the molecular profiles of ICC, KRAS mutation status was assessed in resected specimens. For the assessment of glucose uptake, we observed the expression of glucose transporter-1 (GLUT-1) by immunohistochemistry. The data of 18F-FDG-PET were re-evaluated as follows: maximum standardized uptake value, metabolic tumor volume (MTV), and total lesion glycolysis (TLG). Cut-off values were determined using receiver operating characteristic (ROC) curve analysis. Cumulative overall survival (OS) was analyzed using the Kaplan-Meier analysis. RESULTS: Overall, 16 (32.0%) patients had mutations in KRAS. Patients with mutated KRAS exhibited shorter OS than those with wild-type KRAS (5-year OS, 0% vs. 35.1%, P < 0.001). GLUT-1 expression was significantly higher in tumors with mutated KRAS than in tumors with wild-type KRAS (median, 4.0 vs. 1.0, P < 0.001). Survival was significantly different when stratified by expression of GLUT-1 (5-year OS, 0% vs. 46.5%, P <0.001). Among the 18F-FDG-PET parameters, the MTV and TLG were significantly higher in the mutated KRAS group than in the wild-type KRAS group (P = 0.013 and P = 0.026, respectively). ROC curve analysis revealed a cut-off value of 38 for the MTV, with the highest accuracy (area under the curve = 0.789; 95% confidence interval, 0.581-0.902) for predicting KRAS mutation. This cut-off value permitted stratification of OS (high vs. low: 5-year OS, 13.1% vs. 36.7%, P = 0.008). CONCLUSIONS: High MTV is associated with KRAS mutation and poor postoperative outcomes in patients with ICC, suggesting that the MTV of ICC measured by 18F-FDG-PET may provide useful information for tumor molecular profiles and prognosis.


Assuntos
Colangiocarcinoma/diagnóstico por imagem , Colangiocarcinoma/genética , Fluordesoxiglucose F18/química , Mutação/genética , Tomografia por Emissão de Pósitrons , Cuidados Pré-Operatórios , Proteínas Proto-Oncogênicas p21(ras)/genética , Carga Tumoral , Adulto , Idoso , Idoso de 80 Anos ou mais , Colangiocarcinoma/cirurgia , Feminino , Transportador de Glucose Tipo 1/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Análise de Sobrevida
17.
Cancer Sci ; 108(5): 910-917, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28207963

RESUMO

Nardilysin (NRDC) is a metalloendopeptidase of the M16 family. We previously showed that NRDC activates inflammatory cytokine signaling, including interleukin-6-signal transducer and activator of transcription 3 (STAT3) signaling. NRDC has been implicated in the promotion of breast, gastric and esophageal cancer, as well as the development of liver fibrosis. In this study, we investigated the role of NRDC in the promotion of hepatocellular carcinoma (HCC), both clinically and experimentally. We found that NRDC expression was upregulated threefold in HCC tissue compared to the adjacent non-tumor liver tissue, which was confirmed by immunohistochemistry and western blotting. We also found that high serum NRDC was associated with large tumor size (>3 cm, P = 0.016) and poor prognosis after hepatectomy (median survival time 32.0 vs 73.9 months, P = 0.003) in patients with hepatitis C (n = 120). Diethylnitrosamine-induced hepatocarcinogenesis was suppressed in heterozygous NRDC-deficient mice compared to their wild-type littermates. Gene silencing of NRDC with miRNA diminished the growth of Huh-7 and Hep3B spheroids in vitro. Notably, phosphorylation of STAT3 was decreased in NRDC-depleted Huh-7 spheroids compared to control spheroids. The effect of a STAT3 inhibitor (S3I-201) on the growth of Huh-7 spheroids was reduced in NRDC-depleted cells relative to controls. Our results show that NRDC is a promising prognostic marker for HCC in patients with hepatitis C, and that NRDC promotes tumor growth through activation of STAT3.


Assuntos
Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Metaloendopeptidases/metabolismo , Fator de Transcrição STAT3/metabolismo , Transdução de Sinais/fisiologia , Animais , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Humanos , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/metabolismo , Fosforilação/fisiologia , Prognóstico , Regulação para Cima/fisiologia
18.
Cancer Immunol Immunother ; 66(9): 1131-1142, 2017 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-28432397

RESUMO

Neuropilin-1 (NRP-1)-expressing macrophages are engaged in antitumor immune functions via various mechanisms. In this study, we investigated the role of NRP-1 on macrophages in antibody-mediated tumoricidal activity. Treatment of macrophages with NRP-1 knockdown or an anti-NRP-1-neutralizing antibody significantly suppressed antibody-dependent cellular cytotoxicity and modulated cytokine secretion from macrophages in vitro. Furthermore, in vivo studies using a humanized mouse model bearing human epidermal growth factor receptor-2 (HER2)-positive breast cancer xenografts showed that antibody-mediated antitumor activity and tumor infiltration of CD4+ T lymphocytes were significantly downregulated when peripheral blood mononuclear cells in which NRP-1 was knocked down were co-administered with an anti-HER2 antibody. These results revealed that NRP-1 expressed on macrophages plays an important role in antibody-mediated antitumor immunity. Taken together, the induction of NRP-1 on macrophages may be a therapeutic indicator for antibody treatments that exert antibody-dependent cellular cytotoxicity activity, although further studies are needed in order to support this hypothesis.


Assuntos
Anticorpos Monoclonais/imunologia , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Neoplasias da Mama/imunologia , Macrófagos/metabolismo , Neuropilina-1/imunologia , Animais , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Humanos , Camundongos
19.
Hepatology ; 63(4): 1325-39, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26248612

RESUMO

UNLABELLED: Hepatic stellate cells (HSCs) constitute the liver sinusoid with Kupffer cells and liver sinusoidal endothelial cells. While the sinusoid functions as the gateway to liver inflammation, whether HSCs contribute to liver inflammation and, if so, how they exert such functions remain elusive. Here, we found that mouse as well as human HSCs expressed DP1 receptor for prostaglandin D2 selectively in the liver. Pharmacological stimulation of DP1 by BW245C, a DP1-selective agonist, suppressed the activation of cultured HSCs by tumor necrosis factor-α at least in part through down-regulation of nuclear factor kappa-light-chain-enhancer of activated B cells signaling and inhibition of c-Jun N-terminal kinase phosphorylation. DP1 deficiency or BW245C administration in mice significantly enhanced or suppressed concanavalin A (ConA)-induced hepatitis, respectively. ConA injection induced tumor necrosis factor-α and interferon-γ expression in the sinusoid, which was suppressed by administration of BW245C. Coculture of spleen cells and liver nonparenchymal cells showed that ConA first activated spleen cells and that this activation led to activation of nonparenchymal cells to secondarily produce tumor necrosis factor-α and interferon-γ. Microarray analysis revealed ConA-induced expression of endothelin-1, tissue factor, and chemokines in the liver and inducible nitric oxide synthase in hepatocytes, resulting in flow stagnation, leukocyte adherence and migration to the parenchyma, and hepatocyte death. DP1 stimulation inhibits all these events in the liver. Therefore, HSCs mediate amplification of ConA-induced liver inflammation in the sinusoid, causing direct and indirect hepatocyte injury, and DP1 stimulation inhibits this HSC activation. CONCLUSIONS: HSCs integrate cytokine-mediated inflammatory responses in the sinusoids and relay them to the liver parenchyma, and these HSC actions are inhibited by DP1 stimulation.


Assuntos
Doença Hepática Induzida por Substâncias e Drogas/patologia , Concanavalina A/farmacologia , Células Estreladas do Fígado/metabolismo , Hepatite C/imunologia , Hepatite C/patologia , Animais , Biópsia por Agulha , Estudos de Casos e Controles , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Quimiocinas/efeitos dos fármacos , Quimiocinas/metabolismo , Modelos Animais de Doenças , Feminino , Hepatite C/fisiopatologia , Humanos , Imuno-Histoquímica , Interferon gama/metabolismo , Células de Kupffer/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação/efeitos dos fármacos , Fosforilação/fisiologia , Distribuição Aleatória , Reação em Cadeia da Polimerase em Tempo Real/métodos , Valores de Referência , Sensibilidade e Especificidade , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo
20.
Proc Natl Acad Sci U S A ; 111(32): E3297-305, 2014 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-25074909

RESUMO

Hepatic myofibroblasts are activated in response to chronic liver injury of any etiology to produce a fibrous scar. Despite extensive studies, the origin of myofibroblasts in different types of fibrotic liver diseases is unresolved. To identify distinct populations of myofibroblasts and quantify their contribution to hepatic fibrosis of two different etiologies, collagen-α1(I)-GFP mice were subjected to hepatotoxic (carbon tetrachloride; CCl4) or cholestatic (bile duct ligation; BDL) liver injury. All myofibroblasts were purified by flow cytometry of GFP(+) cells and then different subsets identified by phenotyping. Liver resident activated hepatic stellate cells (aHSCs) and activated portal fibroblasts (aPFs) are the major source (>95%) of fibrogenic myofibroblasts in these models of liver fibrosis in mice. As previously reported using other methodologies, hepatic stellate cells (HSCs) are the major source of myofibroblasts (>87%) in CCl4 liver injury. However, aPFs are a major source of myofibroblasts in cholestatic liver injury, contributing >70% of myofibroblasts at the onset of injury (5 d BDL). The relative contribution of aPFs decreases with progressive injury, as HSCs become activated and contribute to the myofibroblast population (14 and 20 d BDL). Unlike aHSCs, aPFs respond to stimulation with taurocholic acid and IL-25 by induction of collagen-α1(I) and IL-13, respectively. Furthermore, BDL-activated PFs express high levels of collagen type I and provide stimulatory signals to HSCs. Gene expression analysis identified several novel markers of aPFs, including a mesothelial-specific marker mesothelin. PFs may play a critical role in the pathogenesis of cholestatic liver fibrosis and, therefore, serve as an attractive target for antifibrotic therapy.


Assuntos
Cirrose Hepática/patologia , Fígado/patologia , Miofibroblastos/patologia , Animais , Tetracloreto de Carbono/toxicidade , Doença Hepática Induzida por Substâncias e Drogas/genética , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Colestase/complicações , Colágeno Tipo I/genética , Colágeno Tipo I/metabolismo , Cadeia alfa 1 do Colágeno Tipo I , Modelos Animais de Doenças , Proteínas Ligadas por GPI/metabolismo , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Fígado/metabolismo , Cirrose Hepática/etiologia , Cirrose Hepática/metabolismo , Mesotelina , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Miofibroblastos/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Vitamina A/metabolismo
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