RESUMO
Industrial production of food for animals and humans needs increasing amounts of pesticides, especially of organophosphates, which are now easily available worldwide. More than 3 million cases of acute severe poisoning are estimated to occur worldwide every year, and even more cases remain unreported, while 200,000-350,000 incidentally or intentionally poisoned people die every year. Diagnostic and therapeutic procedures in organophosphate poisoning have, however, remained unchanged. In addition to several neurologic symptoms (miosis, fasciculations), hypersecretion of salivary, bronchial, and sweat glands, vomiting, diarrhea, and loss of urine rapidly induce dehydration, hypovolemia, loss of conscience and respiratory distress. Within hours, signs of acidosis due to systemic hypoxia can be observed at first laboratory investigation after hospitalization. While determination of serum-cholinesterase does not have any diagnostic value, it has been established that hypoalbuminemia alone or accompanied by an increase in creatinine, lactate, or C-reactive protein serum levels has negative prognostic value. Increased serum levels of C-reactive protein are a sign of systemic ischemia. Protective mechanical ventilation should be avoided, if possible. In fact, acute respiratory distress syndrome characterized by congestion and increased weight of the lung, accompanied by heart failure, may become the cause of death. As the excess of acetylcholine at the neuronal level can persist for weeks until enough newly, locally synthesized acetylcholinesterase becomes available (the value of oximes in reducing this time is still under debate), after atropine administration, intravenous albumin and fluid infusion should be the first therapeutic interventions to reestablish normal blood volume and normal tissue oxygenation, avoiding death by cardiac arrest.
Assuntos
Insuficiência Cardíaca , Intoxicação por Organofosfatos , Intoxicação , Síndrome do Desconforto Respiratório , Animais , Humanos , Intoxicação por Organofosfatos/diagnóstico , Acetilcolinesterase , Proteína C-Reativa , Causas de Morte , Síndrome do Desconforto Respiratório/etiologia , Síndrome do Desconforto Respiratório/terapia , Intoxicação/diagnóstico , Intoxicação/tratamento farmacológicoRESUMO
Hypercoagulation is one of the major risk factors for ICU treatment, mechanical ventilation, and death in critically ill patients infected with SARS-CoV-2. At the same time, hypoalbuminemia is one risk factor in such patients, independent of age and comorbidities. Especially in patients with severe SARS-CoV-2-infection, albumin infusion may be essential to improve hemodynamics and to reduce the plasma level of the main marker of thromboembolism, namely, the D-dimer plasma level, as suggested by a recent report. Albumin is responsible for 80% of the oncotic pressure in the vessels. This is necessary to keep enough water within the systemic circulatory system and for the maintenance of sufficient blood pressure, as well as for sufficient blood supply for vital organs like the brain, lungs, heart, and kidney. The liver reacts to a decrease in oncotic pressure with an increase in albumin synthesis. This is normally possible through the use of amino acids from the proteins introduced with the nutrients reaching the portal blood. If these are not sufficiently provided with the diet, amino acids are delivered to the liver from muscular proteins by systemic circulation. The liver is also the source of coagulation proteins, such as fibrinogen, fibronectin, and most of the v WF VIII, which are physiological components of the extracellular matrix of the vessel wall. While albumin is the main negative acute-phase protein, fibrinogen, fibronectin, and v WF VIII are positive acute-phase proteins. Acute illnesses cause the activation of defense mechanisms (acute-phase reaction) that may lead to an increase of fibrinolysis and an increase of plasma level of fibrinogen breakdown products, mainly fibrin and D-dimer. The measurement of the plasma level of the D-dimer has been used as a marker for venous thromboembolism, where a fourfold increase of the D-dimer plasma level was used as a negative prognostic marker in critically ill SARS-CoV-2 hospitalized patients. Increased fibrinolysis can take place in ischemic peripheral sites, where the mentioned coagulation proteins can become part of the provisional clot (e.g., in the lungs). Although critically ill SARS-CoV-2-infected patients are considered septic shock patients, albumin infusions have not been considered for hemodynamic resuscitation and as anticoagulants. The role of coagulation factors as provisional components of the extracellular matrix in case of generalized peripheral ischemia due to hypoalbuminemia and hypovolemia is discussed in this review.
Assuntos
Albuminas/administração & dosagem , COVID-19/terapia , Hemodiluição/métodos , Anticoagulantes/administração & dosagem , Coagulação Sanguínea/efeitos dos fármacos , COVID-19/sangue , COVID-19/metabolismo , Estado Terminal/terapia , Fibrinólise/efeitos dos fármacos , Humanos , SARS-CoV-2/isolamento & purificação , TromboelastografiaRESUMO
The mechanisms of radiation-induced liver damage are poorly understood. We investigated if tumour necrosis factor (TNF)-α acts synergistically with irradiation, and how its activity is influenced by platelet endothelial cell adhesion molecule-1 (PECAM-1). We studied murine models of selective single-dose (25 Gy) liver irradiation with and without TNF-α application (2 µg/mouse; i.p.). In serum of wild-type (wt)-mice, irradiation induced a mild increase in hepatic damage marker aspartate aminotransferase (AST) in comparison to sham-irradiated controls. AST levels further increased in mice treated with both irradiation and TNF-α. Accordingly, elevated numbers of leucocytes and increased expression of the macrophage marker CD68 were observed in the liver of these mice. In parallel to hepatic damage, a consecutive decrease in expression of hepatic PECAM-1 was found in mice that received radiation or TNF-α treatment alone. The combination of radiation and TNF-α induced an additional significant decline of PECAM-1. Furthermore, increased expression of hepatic lipocalin-2 (LCN-2), a hepatoprotective protein, was detected at mRNA and protein levels after irradiation or TNF-α treatment alone and the combination of both. Signal transducer and activator of transcription-3 (STAT-3) seems to be involved in the signalling cascade. To study the involvement of PECAM-1 in hepatic damage more deeply, the liver of both wt- and PECAM-1-knock-out-mice were selectively irradiated (25 Gy). Thereby, ko-mice showed higher liver damage as revealed by elevated AST levels, but also increased hepatoprotective LCN-2 expression. Our studies show that TNF-α has a pivotal role in radiation-induced hepatic damage. It acts in concert with irradiation and its activity is modulated by PECAM-1, which mediates pro- and anti-inflammatory signalling.
Assuntos
Fígado/metabolismo , Fígado/patologia , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismo , Radiação Ionizante , Fator de Necrose Tumoral alfa/toxicidade , Animais , Antígenos CD/metabolismo , Antígenos de Diferenciação Mielomonocítica/metabolismo , Aspartato Aminotransferases/sangue , Cinética , Leucócitos/metabolismo , Lipocalina-2/metabolismo , Fígado/efeitos da radiação , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fosforilação/efeitos da radiação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Fator de Transcrição STAT3/metabolismoRESUMO
BACKGROUND: After orthotopic liver transplantation (OLT) for hepatocellular carcinoma (HCC), recurrent HCC mostly develops within 2 years. All cases of de novo HCC described so far occurred later than 2 years after OLT. Prevention of post-transplantation HCC has usually been tried to achieve by curing or controlling recurrent liver disease. This has been rationale for treatment with interferon (IFN)/ribavirin of HCV-recurrence in patients after OLT, transplanted for advanced HCV-induced liver disease and/or HCC. The availability of new and more efficient drugs has improved chances also for previously difficult-to-treat HCV-positive patients. CASE PRESENTATION: A 75 year-old male patient who had undergone OLT for decompensated HCV-cirrhosis in 2009, and bilio-digestive surgery in 2011 under tracrolimus (0.5 mg/day) and prednisone (5 mg/day) immunosuppressive therapy, started to receive antiviral treatment for recurrent HCV-infection of graft with 200 mg/day ribavirin in combination with ledipasvir and sofosbuvir by the end of October 2015. Because of multiple side effects (anemia, asthenia, infections, and reduction of kidney functions - palliated by treatment with erythropoietin), treatment was stopped after 16 weeks. At the third control, a minimal increase in alpha-fetoprotein (AFP) serum level to 10 µg/L was measured 8 months after therapy, whereas both liver sonography and serum transaminases were normal. The patient's general condition; however, remained poor, and a magnetic resonance imaging (MRI) of abdomen was performed 2 months later. A nodule of 3 cm in diameter with a pseudocapsule was found centrally in the liver. The patient had to be hospitalized for recurrent infections of the lung, overt ascites and peritonitis. Rapid tumor growth (10 cm) was detected during last stay in hospital (April 2017), concomitant with a rise of AFP-serum levels to 91 µg/L. The family decided to take the patient home, and best supportive care was provided by a general practitioner, local nurses and the patient's dedicated wife until his death. CONCLUSION: Before treating OLT patients with HCV graft reinfection one should not only consider possible advantages of newly effective antiviral-therapies, but also life expectancy and possible side effects (difficult to manage at an outpatient service basis), including severe disadvantages such as the development of HCC.
Assuntos
Antivirais/uso terapêutico , Carcinoma Hepatocelular/complicações , Hepacivirus/efeitos dos fármacos , Hepatite C/tratamento farmacológico , Neoplasias Hepáticas/complicações , Transplante de Fígado/efeitos adversos , Idoso , Carcinoma Hepatocelular/cirurgia , Hepatite C/etiologia , Humanos , Testes de Função Hepática , Neoplasias Hepáticas/cirurgia , Masculino , Fatores de TempoRESUMO
Kupffer cells are professional phagocytes of the liver clearing bacteria from portal blood. Their clearance capacity, however, can be overwhelmed, transforming them into critical mediators of hepatic-injury. We investigated the consequences of selective Kupffer cell-overload by intraperitoneally administering pyrogen-free gadolinium chloride (GdCl3) or Zymosan into rats and into endotoxin-resistant mice (C3H/HeJ). The number of myeloperoxidase-positive (MPOâº) cells increased at 3 h mainly around the portal vessel after both GdCl3 and Zymosan treatment. Simultaneously, GdCl3 administration reduced detectability of ED-1⺠(but not ED-2) cells near the portal vessel. Serum chemokine (C-X-C motif) ligand 1 (CXCL-1), CXCL-2 and chemokine (C-C motif) ligand 2 (CCL-2) showed a peak at 3 h after both treatment regimens although at a higher extent after Zymosan administration. Accordingly, CXCL-1, CXCL-5 and CCL-2 gene expression in the liver was up-regulated after GdCl3 treatment at 3 h. After Zymosan administration a significant up-regulation of CXCL-1, CXCL-2, CXCL-10, CCL-2, CCL-3 and CCL-20 gene expression in liver at 3 h was observed. After Zymosan administration intracellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule 1 (VCAM-1) gene expression was up-regulated in rat liver tissue. In C3H/HeJ mice both treatment regimens up-regulated CCL-2 and ICAM-1 gene expression after 3 h and down-regulated platelet endothelial cell adhesion molecule 1 (PECAM-1) gene expression. In conclusion, phagocytosis overload of Kupffer cells causes induction of several CXC, CC-chemokines, upregulation of "positive" adhesion molecule gene expression, down-regulation of the "negative" adhesion molecule PECAM-1 and a recruitment of neutrophil granulocytes in the portal area of the liver of treated rats and mice mainly in close contact to the liver macrophages.
Assuntos
Moléculas de Adesão Celular/biossíntese , Quimiocinas/biossíntese , Gadolínio/farmacologia , Fígado/metabolismo , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Zimosan/farmacologia , Animais , Células de Kupffer/metabolismo , Células de Kupffer/patologia , Fígado/patologia , Masculino , Camundongos , Neutrófilos/patologia , Ratos , Ratos Wistar , Regulação para Cima/efeitos dos fármacosRESUMO
The role of the liver and the endocrine pancreas in development of hyperinsulinemia in different types of obesity remains unclear. Sedentary rats (160 g) were fed a low-fat-diet (LFD, chow 13% kcal fat), high-fat-diet (HFD, 35% fat), or HFD+ 30% ethanol+ 30% fructose (HF-EFr, 22% fat). Overnight-fasted rats were culled after one, four or eight weeks. Pancreatic and hepatic mRNAs were isolated for subsequent RT-PCR analysis. After eight weeks, body weights increased three-fold in the LFD group, 2.8-fold in the HFD group, and 2.4-fold in the HF-EFr (p < 0.01). HF-EFr-fed rats had the greatest liver weights and consumed less food during Weeks 4-8 (p < 0.05). Hepatic-triglyceride content increased progressively in all groups. At Week 8, HOMA-IR values, fasting serum glucose, C-peptide, and triglycerides levels were significantly increased in LFD-fed rats compared to that at earlier time points. The greatest plasma levels of glucose, triglycerides and leptin were observed in the HF-EFr at Week 8. Gene expression of pancreatic-insulin was significantly greater in the HFD and HF-EFr groups versus the LFD. Nevertheless, insulin: C-peptide ratios and HOMA-IR values were substantially higher in HF-EFr. Hepatic gene-expression of insulin-receptor-substrate-1/2 was downregulated in the HF-EFr. The expression of phospho-ERK-1/2 and inflammatory-mediators were greatest in the HF-EFr-fed rats. Chronic intake of both LFD and HFD induced obesity, MetS, and intrahepatic-fat accumulation. The hyperinsulinemia is the strongest in rats with the lowest body weights, but having the highest liver weights. This accompanies the strongest increase of pancreatic insulin production and the maximal decrease of hepatic insulin signaling, which is possibly secondary to hepatic fat deposition, inflammation and other factors.
Assuntos
Dieta Hiperlipídica , Resistência à Insulina , Insulina/biossíntese , Síndrome Metabólica/complicações , Síndrome Metabólica/metabolismo , Obesidade/complicações , Obesidade/metabolismo , Animais , Glicemia/metabolismo , Modelos Animais de Doenças , Etanol , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Fígado Gorduroso/patologia , Frutose , Homeostase , Metabolismo dos Lipídeos/genética , Fígado/enzimologia , Fígado/patologia , Masculino , Modelos Biológicos , Tamanho do Órgão , Pâncreas/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Sprague-Dawley , Aumento de PesoRESUMO
BACKGROUND: Melanocortin 4 receptor (MC4R) is predominantly recognized to mediate energy metabolism and anti-inflammation through the central nervous system. However, the expression of MC4R has recently been identified in rat liver and was shown to be upregulated during acute phase response. This study aims to investigate potential roles of MC4R in liver regeneration. MATERIALS AND METHODS: Rat partial hepatectomy (PH) was performed, and MC4R expression was analyzed at different time points after resection. Sham-operated animals (SH) served as controls. In vitro primary hepatocytes (HCs) were isolated from normal rat liver and stimulated with α-melanocyte-stimulating hormone (MC4R agonist). Real-time polymerase chain reaction, Western blot, and immunofluorescence staining were applied to detect gene expression. RESULTS: Up to 8 h after PH, hepatic messenger RNA of proinflammatory cytokines interleukin 6 and tumor necrosis factor α reached peak values. Between 8 and 72 h after PH, rat liver regeneration was extremely active as assessed by the regeneration indices labeled by Ki-67. Immunofluorescence staining indicated that MC4R was mostly expressed in hepatocyte nuclear factor 4(+) cells (HCs) and upregulated during rat liver regeneration. Concurrently, the expression of hepatic MC4R protein was significantly higher in PH than in SH animals, and phosphorylated extracellular signal-regulated kinase 1/2 was remarkably increased in PH compared with SH animals (P < 0.05, respectively). In vitro experiments showed that the expression of proliferating cell nuclear antigen was significantly higher in HCs treated with α-melanocyte-stimulating hormone than in control HCs, which was correlated to the increase of phosphorylated extracellular signal-regulated kinase 1/2 and reduction of phosphorylated signal transducer and activator of transcription 3 (P < 0.05, respectively). CONCLUSIONS: MC4R is predominantly expressed in HCs and upregulated during rat liver regeneration. In vitro stimulation of HC MC4R is associated with a modulation of extracellular signal-regulated kinase and signal transducer and activator of transcription 3 pathways regulating liver regeneration.
Assuntos
Hepatectomia , Regeneração Hepática/fisiologia , Receptor Tipo 4 de Melanocortina/metabolismo , Regulação para Cima , Animais , Biomarcadores/metabolismo , Western Blotting , Citocinas/metabolismo , Imunofluorescência , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase em Tempo Real , Transdução de SinaisRESUMO
Single-dose thioacetamide (TAA) administration induces inflammation and acute liver damage. The mechanism of inflammatory cell recruitment in the liver is still unclear. The aim of this study was to examine the sequence and recruitment of inflammatory cells in different liver regions in relation to CXC- and CC-chemokine and cytokine expression during acute liver injury. Single-dose TAA was administered to rats intraperitoneally, and animals were killed at different time points thereafter. Serum and liver tissue were taken and frozen immediately. Tissue was used for immunostaining cryostat sections, RNA, and protein extraction. RT-PCR and western blotting were performed for RNA and protein analysis, respectively. An early increase (3 h) in CXCL8/IL-8 levels was measured followed by a marked release in MCP1/CCL2 (24 h) serum levels after TAA administration compared with controls. Similarly, an early increase in specific RNA of hepatic chemokines CXCL1/KC and CXCL8/IL-8 was found at 3 h, followed by an upregulation of CXCL5/LIX (6 h), CXCL2/MIP-2 (12 h), and MCP1/CCL2 gene expression at 24-48 h. Further, an induction of pro-inflammatory cytokines IFN-γ and IL-1ß followed by IL-6 and TNF-α was observed with a maximum at 12 h. The magnitude of increase in gene expression of TNF-α and MCP1/CCL2 was the highest among all cytokines and chemokines, respectively. By means of immunohistochemistry, an early (12-24 h) increase in the number of only neutrophil granulocytes (NGs) attached to and around portal vessel walls was observed, followed by increased numbers of mononuclear phagocytes (24-48 h) along the sinusoids. Treatment of the human monocytic cell line U-937 with TNF-α increased the gene expression of CXCL1/KC, CXCL8/IL-8, and MCP1/CCL2. Conversely, adding of infliximab (IFX) to the culture medium inhibited this upregulation significantly. In conclusion, single-dose TAA administration induces a sequence of events with a defined upregulation of gene expression of inflammatory chemokines and cytokines and a transient accumulation of NGs within the portal area and macrophages along the sinusoids throughout the liver. Periportal inflammation seems to precede hepatocellular damage.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Quimiocinas/metabolismo , Citocinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Tioacetamida/toxicidade , Análise de Variância , Animais , Anticorpos Monoclonais , Western Blotting , Quimiocina CCL2/sangue , Primers do DNA/genética , Imuno-Histoquímica , Infliximab , Fagócitos/metabolismo , Ratos , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Tioacetamida/administração & dosagem , Fatores de TempoRESUMO
BACKGROUND: Little is known about the factors that predict for gastrointestinal stromal tumor (GIST) recurrence in patients treated with adjuvant imatinib. METHODS: Risk factors for GIST recurrence were identified, and 2 risk stratification scores were developed using the database of the Scandinavian Sarcoma Group (SSG) XVIII trial, where 358 patients with high-risk GIST with no overt metastases were randomly assigned to adjuvant imatinib 400 mg/day either for 12 or 36 months after surgery. The findings were validated in the imatinib arm of the American College of Surgeons Oncology Group Z9001 trial, where 359 patients with GIST were randomized to receive imatinib and 354 were to receive placebo for 12 months. RESULTS: Five factors (high tumor mitotic count, nongastric location, large size, rupture, and adjuvant imatinib for 12 months) were independently associated with unfavorable recurrence-free survival (RFS) in a multivariable analysis in the SSGXVIII cohort. A risk score based on these 5 factors had a concordance index with GIST recurrence of 78.9%. When a simpler score consisting of the 2 strongest predictive factors (mitotic count and tumor site) was devised, the groups with the lowest, intermediate high, and the highest risk had 5-year RFS of 76.7%, 47.5%, and 8.4%, respectively. Both scores were strongly associated with RFS in the validation cohort (P < .001 for each comparison). CONCLUSIONS: The scores generated were effective in stratifying the risk of GIST recurrence in patient populations treated with adjuvant imatinib. Patients with nongastric GIST with a high mitotic count are at a particularly high risk for recurrence.
Assuntos
Antineoplásicos/administração & dosagem , Benzamidas/administração & dosagem , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Piperazinas/administração & dosagem , Pirimidinas/administração & dosagem , Quimioterapia Adjuvante , Esquema de Medicação , Feminino , Tumores do Estroma Gastrointestinal/patologia , Tumores do Estroma Gastrointestinal/cirurgia , Humanos , Mesilato de Imatinib , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Fatores de RiscoRESUMO
BACKGROUND: Plasma and hepatic lipid abnormalities are frequent in hepatitis C infected individuals. METHODS: Plasma lipid and medical records profiles were prospectively obtained in 130 consecutive individuals seen by a single hepatologist in a university liver disease clinic. The relationships between viral load, genotype, plasma lipid fractions, HDL, LDL particle number and particle size were examined. RESULTS: Of 130 individuals studied, 74 had hepatitis C while 15 had NAFLD/NASH and 30 had alcohol related liver disease. The LDL particle number and LDL-C levels did not differ between those with and without hepatitis C although the number of small LDL particles was greater in those with hepatitis C infection. The HDL-C and total cholesterol levels were greater in those without hepatitis C than those with hepatitis C (P = 0.009). In contrast, the serum triglyceride level was greater in the hepatitis C viral group (P = 0.013). Importantly, the hepatitis C viral load regardless of the genotype correlated directly with the triglyceride and VLDL levels with r values of 0.73 and 0.84, respectively. CONCLUSIONS: There are: (1) important differences in lipid classes, number and the size of lipid particles exist between hepatitis C virus infected and noninfected liver disease groups, (2) the serum total triglyceride and the LDL levels correlate significantly with the hepatitis C viral load and, (3) Serum triglyceride level may play an important role in viral replication. These data further suggest that therapies directed at lowering plasma triglyceride levels may enhance the efficacy of current antiviral treatment regimens.
Assuntos
Colesterol/sangue , Fígado Gorduroso/sangue , Hepacivirus/fisiologia , Hepatopatias Alcoólicas/sangue , Triglicerídeos/sangue , Replicação Viral/fisiologia , Adulto , HDL-Colesterol/sangue , LDL-Colesterol/sangue , VLDL-Colesterol/sangue , Comorbidade , Diabetes Mellitus Tipo 2/epidemiologia , Fígado Gorduroso/epidemiologia , Feminino , Genótipo , Hepacivirus/genética , Hepatite C/epidemiologia , Humanos , Hepatopatias Alcoólicas/epidemiologia , Masculino , Pessoa de Meia-Idade , Hepatopatia Gordurosa não Alcoólica , Tamanho da Partícula , Estudos ProspectivosRESUMO
It has been recently shown that the biological effects of erythropoietin (EPO) are not limited to the hematopoietic compartment but, as pleiotropic glycoprotein, this hormone can exert pro-angiogenic and tissue-protective functions also in a wide range of non-hematopoietic organs. The role of EPO and the effective functionality of its receptor in solid tumors are still a controversial point of debate. In the present work we analyzed the gene expression of EPO and its cognate receptor (EpoR) in a rat model of thioacetamide-induced damage and tumor. An analysis of the EPO/EpoR axis was also performed on human cholangiocarcinoma (CC) cell lines. A progressive increase of EPO and EpoR mRNA can already be observed during the fibrotic-cirrhotic development with a peak of expression rising at tumor formation (24.7 ± 9.9-fold increase and 15.5 ± 1.1-fold increase, respectively, for the two genes). Co-localization studies by immunofluorescence revealed hepatocytes in the regenerative cirrhotic nodules (Hep Par-1(+)) and in the dysplastic bile duct cells (CK19(+)) as the major EPO producers in this specific condition. The same cell populations, together with endothelial cells, exhibited an increased expression of EpoR, although all the non-parenchymal cell populations in the liver exhibited modest basal mRNA levels. Challenging human CC cells, Mz-Cha-2, with a combination of EPO and SCF resulted in a synergistic effect on the gene expression of EPO, CyclinD1 and PCNA. This study suggests that the autocrine and paracrine release of endogenous EPO in the microenvironment may contribute to the development and maintenance of the CC possibly in cooperation with other signaling pathways.
Assuntos
Neoplasias dos Ductos Biliares/metabolismo , Colangiocarcinoma/metabolismo , Modelos Animais de Doenças , Eritropoetina/metabolismo , Fígado/metabolismo , Fígado/patologia , Receptores da Eritropoetina/metabolismo , Animais , Neoplasias dos Ductos Biliares/patologia , Colangiocarcinoma/patologia , Eritropoetina/genética , Humanos , Fígado/lesões , Masculino , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores da Eritropoetina/genética , Tioacetamida , Células Tumorais CultivadasRESUMO
The circulating 25-hydroxylated form of vitamin D(3), 25(OH)D, and serum ferritin concentrations have been described to be associated with disease progression in chronic hepatitis C. Both parameters also have been assessed with regard to treatment outcome, however, with divergent results. This study examined both the pre- and posttreatment serum concentrations of 25(OH)D and ferritin in 191 patients infected chronically with hepatitis C virus (HCV) type 1 with regard to liver inflammatory activity (grading), disease progression in terms of fibrosis (staging) and an antiviral treatment outcome. Mean pretreatment serum 25(OH)D and ferritin concentrations were 18 ± 10 ng/ml and 280 ± 225 µg/L, respectively. Multivariate analysis revealed lower pretreatment serum 25(OH)D and higher ferritin concentrations to be significantly related to both severity of inflammatory activity and of fibrotic alterations. Pretreatment serum ferritin concentration, furthermore, unlike 25(OH)D concentration, was found to be associated with a sustained virological response by uni- and multivariate analyses. A sustained virological response was featured by a significant increase in serum 25(OH)D levels (18 ± 10 ng/ml vs. 22 ± 11 ng/ml; P < 0.01), a reduction of serum ferritin concentration (191 ± 156 µg/L vs. 103 ± 63 µg/L; P < 0.001) and a normalization of serum alanine aminotransferase (ALT) and γ-glutamyl-transferase (γ-GT) activities. Taken together, decreased 25(OH)D and increased ferritin serum levels indicate the severity of hepatic inflammation and fibrosis in patients infected chronically with HCV type 1. Elevated ferritin, furthermore, was found to be an independent predictor for standard IFN-based therapy responsiveness.
Assuntos
Biomarcadores/sangue , Calcifediol/sangue , Ferritinas/sangue , Hepacivirus/classificação , Hepatite C Crônica/patologia , Adulto , Idoso , Progressão da Doença , Feminino , Hepacivirus/genética , Hepacivirus/isolamento & purificação , Hepatite C Crônica/virologia , Humanos , Cirrose Hepática/patologia , Masculino , Pessoa de Meia-Idade , Soro/química , Índice de Gravidade de Doença , Resultado do TratamentoRESUMO
BACKGROUND/AIM: IL-6 - IL-1- lipocalin2 (LCN2) - liver irradiation - oxidative stress - TNF-a Lipocalin2 (LCN2) is an acute phase protein. The source of its increased serum level in oxidative stress conditions (ROS) remains still unknown. We prospectively evaluate the serum LCN2 increase after single dose liver irradiation along with hepatic LCN2 gene and protein expression. METHODS: A single dose of 25 Gray was administered percutaneously to the liver of randomly paired rats after a planning CT scan. Male Wistar rats were sacrificed 1, 3, 6, 12, 24 and 48 h after irradiation along with sham-irradiated controls. ELISA, RT-PCR, Western blot and immunofluorescence staining was performed. Furthermore, hepatocytes, myofibroblasts and Kupffer cells were isolated from the liver of healthy rats and irradiated ex-vivo. RESULTS: After liver irradiation, LCN2 serum levels increased significantly up to 2.7 µg/ml within 6 h and stayed elevated over 24 h. LCN2 specific transcripts increased significantly up to 552 ± 109-fold at 24 h after liver irradiation, which was further confirmed at protein level. α2-macroglobulin and hemoxygenase-1 also showed an increase, but the magnitude was less as compared to LCN2. LCN2+ granulocytes were detected within 1 h after irradiation around central and portal fields and remained high during the course of study. Ex-vivo irradiated hepatocytes (2.4 ± 0.6-fold) showed a higher LCN2 gene expression as compared to myofibroblasts and Kupffer cells. IL-1ß treatment further increased LCN2 gene expression in cultured hepatocytes. CONCLUSIONS: Single dose liver irradiation induces a significant increase in LCN2 serum levels, comparable to the induction of acute phase proteins. We suggest LCN2 as marker for the early phase of radiation-induced tissue damage.
Assuntos
Biomarcadores/sangue , Regulação da Expressão Gênica/fisiologia , Lipocalinas/sangue , Fígado/lesões , Fígado/efeitos da radiação , Lesões por Radiação/diagnóstico , Animais , Western Blotting , Primers do DNA/genética , Ensaio de Imunoadsorção Enzimática , Imunofluorescência , Humanos , Interleucina-1/metabolismo , Interleucina-6/metabolismo , Lipocalina-2 , Masculino , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TempoRESUMO
PURPOSE: The duodenum as primary site for gastrointestinal stromal tumors (GISTs) is rare and mitotic rate, tumor size, type of mutation and number of chromosomal aberrations have prognostic implications. METHODS: We analyzed the outcome of 13 patients with duodenal GISTs who underwent surgical tumor resection. Either segmental duodenectomy or pylorus-preserving duodenopancreatectomy was performed. The tumors were histopathologically examined and the risk of progression was assessed based on tumor size and mitotic count. Additionally, mutation analysis of the KIT and PDGFRA receptor tyrosine kinase genes and comparative genomic hybridization (CGH) were performed in all cases. RESULTS: Eight patients underwent segmental duodenectomy and five patients were treated with pylorus-preserving duodenopancreatectomy. None of the five GISTs with low or no risk for malignancy according to the Miettinen classification developed tumor progress. In contrast, five of eight cases (62.5%) with high-risk tumors revealed tumor progress, and four of these patients died (50%). The median overall survival for all patients was 66 months, and the median disease-free survival 41 months. The operative procedure and type of mutation did not correlate with long-term survival. CGH analysis displayed -15q in 12/13 tumors, and -1p in 11/13 cases as characteristic chromosomal aberrations for intestinal origin. Notably, -22q was present in three of four cases with tumor progress. CONCLUSIONS: Both segmental duodenectomy and pylorus-preserving duodenopancreatectomy are appropriate options to treat duodenal GIST and should be implemented depending on resectability and the patient's performing state. The Miettinen classification and CGH findings correlate with the clinical course.
Assuntos
Duodeno/patologia , Tumores do Estroma Gastrointestinal/patologia , Idoso , Aberrações Cromossômicas , Hibridização Genômica Comparativa , Análise Mutacional de DNA , Procedimentos Cirúrgicos do Sistema Digestório/efeitos adversos , Intervalo Livre de Doença , Duodeno/cirurgia , Éxons/genética , Feminino , Seguimentos , Tumores do Estroma Gastrointestinal/cirurgia , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Complicações Pós-Operatórias/etiologia , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genéticaRESUMO
The liver is considered a radiosensitive organ. However, in rats, high single-dose irradiation (HDI) showed only mild effects. Consequences of fractionated irradiation (FI) in such an animal model have not been studied so far. Rats were exposed to selective liver FI (total dose 60 Gy, 2 Gy/day) or HDI (25 Gy) and were killed three months after the end of irradiation. To study acute effects, HDI-treated rats were additionally killed at several time points between 1 and 48 h. Three months after irradiation, no differences between FI and HDI treatment were found for macroscopically detectable small "scars" on the liver surface and for an increased number of neutrophil granulocytes distributed in the portal fields and through the liver parenchyma. As well, no changes in HE-stained tissues or clear signs of fibrosis were found around the portal vessels. Differences were seen for the number of bile ducts being increased in FI- but not in HDI-treated livers. Serum levels indicative of liver damage were determined for alkaline phosphatase (AP), aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyltransferase (γGT) and lactate dehydrogenase (LDH). A significant increase of AP was detected only after FI while HDI led to the significant increases of AST and LDH serum levels. By performing RT-PCR, we detected up-regulation of matrix metalloproteinases, MMP-2, MMP-9, MMP-14, and of their inhibitors, TIMP-1, TIMP-2 and TIMP-3, shortly after HDI, but not at 3 month after FI or HDI. Overall, we saw punctual differences after FI and HDI, and a diffuse formation of small scars at the liver surface. Lack of "provisional clot"-formation and absence of recruitment of mononuclear phagocytes could be one explanation for scar formation as incomplete repair response to irradiation.
Assuntos
Fígado/efeitos da radiação , Radiação Ionizante , Alanina Transaminase/sangue , Fosfatase Alcalina/sangue , Animais , Aspartato Aminotransferases/sangue , L-Lactato Desidrogenase/sangue , Contagem de Leucócitos , Fígado/patologia , Masculino , Tamanho do Órgão/efeitos da radiação , Doses de Radiação , Ratos , Ratos Wistar , Tempo , Inibidor Tecidual de Metaloproteinase-1/genética , Inibidor Tecidual de Metaloproteinase-1/metabolismo , Inibidor Tecidual de Metaloproteinase-2/genética , Inibidor Tecidual de Metaloproteinase-2/metabolismo , gama-Glutamiltransferase/sangueRESUMO
Chronic inflammatory bowel diseases can be successfully treated with antibodies against the acute phase mediator TNF-α. The process of activation and of extravasation of inflammatory cells from the blood into the 'stressed' tissue site is controlled by cytokines and chemokines, which attract leukocytes and by adhesion molecules, which mediate their attachment and transmigration toward the affected cell(s). The changes in the gene expression of adhesion molecules taking place in those cells before attachment have been less investigated. Changes of PECAM-1, ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1) gene expression were studied in phytohaemagglutinin (PHA)- and lipolysaccharide (LPS)-treated human peripheral blood leukocytes (PBLs), granulocytes and the human monocyte cell line U-937. Cells were treated either with PHA or with LPS in the presence or absence of infliximab and incubated with TNF-α, IFN-γ and/or transforming growth factor beta (TGF-ß) and treated as above. Activation of PBLs by PHA or LPS treatment triggered a sharp upregulation of ICAM-1, VCAM-1 gene expression and a time-dependent downregulation of PECAM-1 gene expression reaching a minimum 4 h from start of the experiment. The anti-TNF-α antibody infliximab, by neutralizing TNF-α and IFN-γ production, completely reversed PECAM-1 mRNA downregulation and ICAM-1 and VCAM-1 upregulation. Immunostaining of PBLs cytospins with antibodies against PECAM-1 and ICAM-1 confirmed RT-PCR and western blot results. PBLs IFN-γ or TNF-α treatment downregulated PECAM-1 in parallel with the upregulation of ICAM-1 and VCAM-1 gene expression, whereas TGF-ß upregulated PECAM-1- and downregulated ICAM-1 and VCAM-1 gene expression counteracting the effect of TNF-α or IFN-γ. Similar results were obtained in human U937 cells and in granulocyte cultures by TNF-α or IFN-γ treatment. Taken together, these results suggest that infliximab, blocking TNF-α and IFN-γ production, exerts its anti-inflammatory effect through inhibiting downregulation of PECAM-1 gene expression and upregulation of ICAM-1 and VCAM-1 expression in leukocytes of the peripheral blood. These results also suggest that TGF-ß may thus be of therapeutic importance as an anti-inflammatory agent.
Assuntos
Anticorpos Monoclonais/farmacologia , Fármacos Gastrointestinais/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Leucócitos/efeitos dos fármacos , Modelos Teóricos , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Sequência de Bases , Western Blotting , Células Cultivadas , Citocinas/metabolismo , Primers do DNA , Humanos , Técnicas In Vitro , Infliximab , Molécula 1 de Adesão Intercelular/genética , Leucócitos/metabolismo , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Molécula 1 de Adesão de Célula Vascular/genéticaRESUMO
Liver is the central organ of iron metabolism. During acute-phase-response (APR), serum iron concentration rapidly decreases. The current study aimed to compare expression and localization of iron transport protein ferroportin-1 (Fpn-1) and of other iron import proteins after experimental tissue damage induced by injecting turpentine oil in the hind limbs of rats and mice. Serum and spleen iron concentration decreased with an increase in total liver, cytoplasmic and nuclear iron concentration. In liver, mRNA amount of Fpn-1, Fpn-1a, Fpn-1b, HFE, hemojuvelin (HJV) and hephaestin (heph) genes showed a rapid decrease. Hepcidin, divalent metal transporter-1 (DMT-1), transferrin (Tf) and Tf-receptor-1 (TfR1), TfR-2 (TfR2) gene expression was increased. Western blot analysis of liver tissue lysate confirmed the changes observed at mRNA level. In spleen, a rapid decrease in gene expression of Fpn-1, Fpn-1a, Fpn-1b, DMT-1, Tf, TfR1 and TfR2, and an increase in hepcidin was observed. Immunohistochemistry of DMT-1 and TfR2 were mainly detected in the nucleus of rat liver and spleen, whereas TfR1 was clearly localized in the plasma membrane. Fpn-1 was mostly found in the nuclei of liver cells, whereas in spleen, the protein was mainly detected in the cell membrane. Western blot analysis of liver fractions confirmed immunohistochemical results. In livers of wild-type mice, gene expression of Fpn-1, Fpn-1a and Fpn-1b was downregulated, whereas hepcidin gene expression was increased. In contrast, these changes were less pronounced in IL-6ko-mice. Cytokine (IL-6, IL-1b and TNF-a) treatment of rat hepatocytes showed a downregulation of Fpn-1, Fpn-1a and Fpn-1b, and upregulation of hepcidin gene expression. Moreover, western blot analysis of cell lysate of IL-6-treated hepatocytes detected, as expected, an increase of a2-macroglobulin (positive acute-phase protein), whereas albumin (negative acute-phase protein) and Fpn-1 were downregulated. Our results demonstrate that liver behaves as a 'sponge' for iron under acute-phase conditions, and Fpn-1 behaves as a negative acute-phase protein in rat hepatocytes mainly, but not exclusively, because of the effect of IL-6. These changes could explain iron retention in the cytoplasm and in the nucleus of hepatocytes during APR.
Assuntos
Proteínas de Fase Aguda/metabolismo , Reação de Fase Aguda/metabolismo , Proteínas de Transporte de Cátions/metabolismo , Proteínas de Ligação ao Ferro/metabolismo , Proteínas Reguladoras de Ferro/metabolismo , Fígado/metabolismo , Reação de Fase Aguda/induzido quimicamente , Animais , Proteínas de Transporte de Cátions/genética , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Células Cultivadas , Citoplasma/efeitos dos fármacos , Citoplasma/metabolismo , Citoplasma/patologia , Modelos Animais de Doenças , Expressão Gênica , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/patologia , Interleucina-6/deficiência , Interleucina-6/farmacologia , Ferro/análise , Ferro/metabolismo , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , RNA Mensageiro/metabolismo , Ratos , Ratos Endogâmicos , Terebintina/toxicidadeRESUMO
Interferon regulatory factor-1 (IRF-1), a transcription regulator involved both in inducing and in mediating the effects of interferon, is encoded by a highly polymorphic gene in different ethnic populations. Some of these genetic variations have been described to be associated to disease traits in hepatitis C virus and in human immunodeficiency virus infection, including one single-nucleotide polymorphism rs2549009 within the promoter region. This study aimed at investigating the functional relevance of rs2549009 on IRF-1 transcriptional activity in peripheral blood mononuclear cells in its natural genomic environment. Haplotype-specific chromatin immunoprecipitation using antibodies directed against both the transcriptionally inactive and active RNA polymerase II (RNAPII) and allele-specific transcript quantification techniques were applied to ex vivo-derived samples from healthy heterozygous donors. Inactive serine 5 phosphorylated RNAPII was found to be preferentially bound to the rs2549009 A allele in all donors investigated. Active serine 2 phosphorylated (ser2-P) RNAPII, in contrast, was found to be precipitable, depending on the donor, preferentially either with the A or the G promoter variants or without any preference. The ratio of rs2549009 A/G promoter variants engaged by ser2-P RNAPII was closely related to the relative frequency of the respective IRF-1 transcripts, and relative allelic expression was found to be associated to total IRF-1 gene expression. These results provide evidence for a bidirectional IRF-1 gene expression imbalance that appears not to be solely controlled by rs2549009 in cis and may rely on a yet unidentified variant or haplotype or on environmental control in trans.
Assuntos
Fator Regulador 1 de Interferon/genética , Fator Regulador 1 de Interferon/metabolismo , Polimorfismo de Nucleotídeo Único/genética , Transcrição Gênica , Adulto , Alelos , Imunoprecipitação da Cromatina , Feminino , Expressão Gênica , Variação Genética , Haplótipos , Humanos , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Fosfosserina/metabolismo , Reação em Cadeia da Polimerase , Regiões Promotoras Genéticas , RNA Polimerase II/imunologia , RNA Polimerase II/metabolismo , Elementos Reguladores de TranscriçãoRESUMO
MCRs are known to be expressed predominantly in the brain where they mediate metabolic and anti-inflammatory functions. Leptin plays an important role in appetite and energy regulation via signaling through melanocortin receptors (MCRs) in the brain. As serum levels of MCR ligands are elevated in a clinical situation [acute-phase response (APR)] to tissue damage, where the liver is responsible for the metabolic changes, we studied hepatic gene expression of MCRs in a model of muscle tissue damage induced by turpentine oil (TO) injection in rats. A significant increase in gene expression of all five MCRs (MC4R was the highest) in liver at the RNA and protein level was detected after TO injection. A similar pattern of increase was also found in the brain. Immunohistology showed MC4R in the cytoplasm, but also in the nucleus of parenchymal and non-parenchymal liver cells, whereas MC3R-positivity was mainly cytoplasmic. A time-dependent migration of MC4R protein from the cytoplasm into the nucleus was observed during APR, in parallel with an increase in α-MSH and leptin serum levels. An increase of MC4R was detected at the protein level in wild-type mice, while such an increase was not observed in IL-6ko mice during APR. Moreover, treatment of isolated liver cells with melanocortin agonists (α-MSH and THIQ) inhibited the endotoxin-induced upregulation of the acute-phase cytokine (IL-6, IL1ß and TNF-α) gene expression in Kupffer cells and of chemokine gene expression in hepatocytes. MCRs are expressed not only in the brain, but also in liver cells and their gene expression in liver and brain tissue is upregulated during APR. Due to the presence of specific ligands in the serum, they may mediate metabolic changes and exert a protective effect on liver cells.
Assuntos
Reação de Fase Aguda/imunologia , Fígado/imunologia , Receptores de Melanocortina/genética , Receptores de Melanocortina/imunologia , Animais , Encéfalo/fisiologia , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Células Hep G2 , Hepatócitos/efeitos dos fármacos , Hepatócitos/imunologia , Humanos , Interleucina-6/genética , Células de Kupffer/efeitos dos fármacos , Células de Kupffer/imunologia , Leptina/sangue , Fígado/citologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ratos , Ratos Wistar , Receptor Tipo 1 de Melanocortina/genética , Receptor Tipo 1 de Melanocortina/imunologia , Receptor Tipo 2 de Melanocortina/genética , Receptor Tipo 2 de Melanocortina/imunologia , Receptor Tipo 3 de Melanocortina/genética , Receptor Tipo 3 de Melanocortina/imunologia , Receptor Tipo 4 de Melanocortina/genética , Receptor Tipo 4 de Melanocortina/imunologia , Tetra-Hidroisoquinolinas/farmacologia , Triazóis/farmacologia , alfa-MSH/sangue , alfa-MSH/farmacologiaRESUMO
It has been suggested that cyclooxygenase-2 (COX-2)-mediated prostaglandin synthesis is associated with liver inflammation and carcinogenesis. The aim of this study is to identify the cellular source of COX-2 expression in different stages, from acute liver injury through liver fibrosis to cholangiocarcinoma (CC). We induced in rats acute and "chronic" liver injury (thioacetamide (TAA) or carbon tetrachloride (CCl(4))) and CC development (TAA) and assessed COX-2 gene expression in normal and damaged liver tissue by RT-PCR of total RNA. The cellular localization of COX-2 protein in liver tissue was analyzed by immunohistochemistry as well as in isolated rat liver cells by Western blotting. The findings were compared with those obtained in human cirrhotic liver tissue. The specificity of the antibodies was tested by 2-DE Western blot and mass spectrometric identification of the positive protein spots. RT-PCR analysis of total RNA revealed an increase of hepatic COX-2 gene expression in acutely as well as "chronically" damaged liver. COX-2-protein was detected in those ED1(+)/ED2(+) cells located in the non-damaged tissue (resident tissue macrophages). In addition COX-2 positivity in inflammatory mononuclear phagocytes (ED1(+)/ED2(-)), which were also present within the tumoral tissue was detected. COX-2 protein was clearly detectable in isolated Kupffer cells as well as (at lower level) in isolated "inflammatory" macrophages. Similar results were obtained in human cirrhotic liver. COX-2 protein is constitutively detectable in liver tissue macrophages. Inflammatory mononuclear phagocytes contribute to the increase of COX-2 gene expression in acute and chronic liver damage induced by different toxins and in the CC microenvironment.