RESUMO
Two decades of research identified more than a dozen clock genes and defined a biochemical feedback mechanism of circadian oscillator function. To identify additional clock genes and modifiers, we conducted a genome-wide small interfering RNA screen in a human cellular clock model. Knockdown of nearly 1000 genes reduced rhythm amplitude. Potent effects on period length or increased amplitude were less frequent; we found hundreds of these and confirmed them in secondary screens. Characterization of a subset of these genes demonstrated a dosage-dependent effect on oscillator function. Protein interaction network analysis showed that dozens of gene products directly or indirectly associate with known clock components. Pathway analysis revealed these genes are overrepresented for components of insulin and hedgehog signaling, the cell cycle, and the folate metabolism. Coupled with data showing many of these pathways are clock regulated, we conclude the clock is interconnected with many aspects of cellular function.
Assuntos
Relógios Biológicos , Ritmo Circadiano , Estudo de Associação Genômica Ampla , Linhagem Celular , Técnicas de Silenciamento de Genes , Humanos , Interferência de RNA , RNA Interferente Pequeno/metabolismoRESUMO
Chagas disease, leishmaniasis and sleeping sickness affect 20 million people worldwide and lead to more than 50,000 deaths annually. The diseases are caused by infection with the kinetoplastid parasites Trypanosoma cruzi, Leishmania spp. and Trypanosoma brucei spp., respectively. These parasites have similar biology and genomic sequence, suggesting that all three diseases could be cured with drugs that modulate the activity of a conserved parasite target. However, no such molecular targets or broad spectrum drugs have been identified to date. Here we describe a selective inhibitor of the kinetoplastid proteasome (GNF6702) with unprecedented in vivo efficacy, which cleared parasites from mice in all three models of infection. GNF6702 inhibits the kinetoplastid proteasome through a non-competitive mechanism, does not inhibit the mammalian proteasome or growth of mammalian cells, and is well-tolerated in mice. Our data provide genetic and chemical validation of the parasite proteasome as a promising therapeutic target for treatment of kinetoplastid infections, and underscore the possibility of developing a single class of drugs for these neglected diseases.
Assuntos
Doença de Chagas/tratamento farmacológico , Kinetoplastida/efeitos dos fármacos , Kinetoplastida/enzimologia , Leishmaniose/tratamento farmacológico , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Inibidores de Proteassoma/farmacologia , Inibidores de Proteassoma/uso terapêutico , Pirimidinas/farmacologia , Triazóis/farmacologia , Tripanossomíase Africana/tratamento farmacológico , Animais , Doença de Chagas/parasitologia , Quimotripsina/antagonistas & inibidores , Quimotripsina/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Concentração Inibidora 50 , Leishmaniose/parasitologia , Camundongos , Estrutura Molecular , Terapia de Alvo Molecular , Inibidores de Proteassoma/efeitos adversos , Inibidores de Proteassoma/classificação , Pirimidinas/efeitos adversos , Pirimidinas/química , Pirimidinas/uso terapêutico , Especificidade da Espécie , Triazóis/efeitos adversos , Triazóis/química , Triazóis/uso terapêutico , Tripanossomíase Africana/parasitologiaRESUMO
The basic way of invasion and metastasis of lung cancer is that the tumor cells shed in the extracellular matrix, invade the basement membrane and the surrounding tissue, infiltrate into blood flow, and then survive and transport via the blood flow. After having been extravasated, migrated and arrested in the distant site, they finally form a metastatic lesion. Some basic mechanisms are required in these steps, such as tumor stem cells, diffusion and activity of tumor cells, escaping from apoptosis, angiogenesis and lymphangiogenesis, infiltration into blood flow, circulation and exudation, and distant metastasis proliferation. A better understanding of the mechanisms of the invasion and metastasis of lung cancer will facilitate the prevention and treatment of lung cancer.
Assuntos
Neoplasias Pulmonares , Apoptose , Humanos , Invasividade Neoplásica , Metástase Neoplásica , Células-Tronco Neoplásicas , Neovascularização PatológicaRESUMO
Allergic bronchopulmonary aspergillosis is one of major pulmonary fungal diseases. Although it is not a rare in clinical settings,the misdiagnosis rate is high and the treatment effectiveness remains unstable. This article reviews the recent advances in the diagnosis and treatment of this disease.
Assuntos
Aspergilose Broncopulmonar Alérgica/diagnóstico , Aspergilose Broncopulmonar Alérgica/terapia , Humanos , Resultado do TratamentoRESUMO
Two CYP51 inhibitors, posaconazole and the ravuconazole prodrug E1224, were recently tested in clinical trials for efficacy in indeterminate Chagas disease. The results from these studies show that both drugs cleared parasites from the blood of infected patients at the end of the treatment but that parasitemia rebounded over the following months. In the current study, we sought to identify a dosing regimen of posaconazole that could permanently clear Trypanosoma cruzi from mice with experimental Chagas disease. Infected mice were treated with posaconazole or benznidazole, an established Chagas disease drug, and parasitological cure was defined as an absence of parasitemia recrudescence after immunosuppression. Twenty-day therapy with benznidazole (10 to 100 mg/kg of body weight/day) resulted in a dose-dependent increase in antiparasitic activity, and the 100-mg/kg regimen effected parasitological cure in all treated mice. In contrast, all mice remained infected after a 25-day treatment with posaconazole at all tested doses (10 to 100 mg/kg/day). Further extension of posaconazole therapy to 40 days resulted in only a marginal improvement of treatment outcome. We also observed similar differences in antiparasitic activity between benznidazole and posaconazole in acute T. cruzi heart infections. While benznidazole induced rapid, dose-dependent reductions in heart parasite burdens, the antiparasitic activity of posaconazole plateaued at low doses (3 to 10 mg/kg/day) despite increasing drug exposure in plasma. These observations are in good agreement with the outcomes of recent phase 2 trials with posaconazole and suggest that the efficacy models combined with the pharmacokinetic analysis employed here will be useful in predicting clinical outcomes of new drug candidates.
Assuntos
Inibidores de 14-alfa Desmetilase/farmacologia , Doença de Chagas/tratamento farmacológico , Nitroimidazóis/farmacologia , Parasitemia/tratamento farmacológico , Triazóis/farmacologia , Tripanossomicidas/farmacologia , Inibidores de 14-alfa Desmetilase/farmacocinética , Administração Oral , Animais , Doença de Chagas/enzimologia , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Ensaios Clínicos Fase II como Assunto , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Esquema de Medicação , Coração/efeitos dos fármacos , Coração/parasitologia , Humanos , Terapia de Imunossupressão , Camundongos , Células NIH 3T3 , Nitroimidazóis/farmacocinética , Parasitemia/enzimologia , Parasitemia/imunologia , Parasitemia/parasitologia , Recidiva , Esterol 14-Desmetilase/metabolismo , Triazóis/farmacocinética , Tripanossomicidas/farmacocinética , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/patogenicidade , Trypanosoma cruzi/fisiologiaRESUMO
OBJECTIVE: To explore evolution rules of phlegm and blood stasis syndrome ( PBSS) in hyperlipidemia and atherosclerosis (AS) using NMR-based metabolic profiling and metabonomic approaches based on formulas corresponding to syndrome. METHODS: Totally 150 SD rats were divided into the normal group, the model group, the Erchen Decoction (ED) group, the Xuefu Zhuyu Decoction (XZD) group, the Lipitor group, 30 in each group. The hyperlipidemia and AS rat model was duplicated by suturing carotid artery, injecting vitamin D3, and feeding with high fat diet. ED and XZD were used as drug probes. Blood samples were withdrawn at week 2, 4, and 8 after modeling. Blood lipids, blood rheology, histopathology and metabolomics were detected and analyzed. Results Results of blood lipids and pathology showed hyperlipidemia and early AS rat models were successfully established. At week 2 after modeling, levels of total cholesterol (TC) and low-density lipoprotein cholesterol (LDL-C) significantly increased, which reached the peak at week 4 and maintained at higher levels at week 8. ED exerted obvious effect in improving TC and LDL-C levels of early models, while XZD could greatly improve levels of TC and LDL-C of late models. Rheological results showed at week 2, there was no significant difference in whole blood viscosity, plasma viscosity, or hematocrit between the model group and the normal group (P > 0.05). At week 4 partial hemorheological indicators (such as plasma viscosity) were abnormal. Till week 8 whole blood viscosity, plasma viscosity, and hematocrit were significantly abnormal (P <0. 05, P < 0.01). As time went by, whole blood viscosity, plasma viscosity, and hematocrit showed gradual increasing tendency in the ED group, while they showed gradual decreasing tendency in the XZD group. Results of metabonomics showed significant difference in spectra of metabolites between the normal group and the model group. As modeling time was prolonged, contents of acetyl glucoprotein and glucose in the model group increased in late stage, which was in. line with results of blood lipids and hemorheology. ED showed more obvious effect in early and mid-term modeling (at week 2 and 4), and increased contents of partial metabolites (such as choline, phosphatidyl choline, glycerophosphocholine), but these changes in the XZD group were consistent with those of the model group. In late modeling (at week 8) XZD showed more obvious effect in improving contents of lactic acid, acetyl glycoprotein, LDL, creatine, choline, and glucose. CONCLUSIONS: ED and XZD not only showed regulatory effects on lipid disorders, but also could improve dysbolism of Chos. In formulas corresponding to syndrome, damp-phlegm was main pathogenesis of hyperlipidema and AS in early and mid stages. Blood stasis syndrome began to occur along with it progressed. Phlegm can result in blood stasis and intermingles with stasis. Phlegm turbidity runs through the whole process.
Assuntos
Aterosclerose/metabolismo , Medicamentos de Ervas Chinesas/uso terapêutico , Metaboloma/fisiologia , Escarro/metabolismo , Animais , Colesterol , LDL-Colesterol , Hemorreologia , Hiperlipidemias , Lipídeos , Imageamento por Ressonância Magnética , Medicina Tradicional Chinesa , Metabolômica , Ratos , Ratos Sprague-DawleyRESUMO
Methylenetetrahydrofolate reductase (MTHFR) has been demonstrated to be involved in carcinogenesis. Increasing individual studies have investigated the role of MTHFR C677T polymorphism in gastric cancer pathogenesis, but with inconsistent findings. The aim of this study was to clarify the potential association of the MTHFR C677T polymorphism with gastric cancer risk by pooling all available data from published case-control studies. We searched the PubMed, Embase, Web of Science, and Wanfang databases for all relevant publications to date. The pooled odds ratio (OR) with corresponding 95 % confidence interval (95% CI) was calculated. Stratified analysis and sensitivity analysis were also carried out to estimate the strength of this association. A total of 25 case-control studies with 6,572 cases and 9,584 controls were retrieved. Overall, the ORs under five contrast models indicated that the MTHFR C677T variant was positively associated with gastric cancer risk (ORT vs. C = 1.21, 95% CI 1.101.34, P(OR) < 0.001; OR(TT vs. CC) = 1.47, 95% CI 1.221.76, P(OR) < 0.001; OR(TC vs. CC) = 1.20, 95% CI 1.03-1.40, P(OR) = 0.022; OR(TT + TC vs. CC) = 1.27, 95% CI 1.10-1.47, P(OR) = 0.001; OR(TT vs. CC + TC) = 1.29, 95% CI 1.15-1.46, P(OR) < 0.001). Stratified analyses according to ethnicity and source of controls further confirmed the significant correlations. The current meta-analysis provides strong evidence that the MTHFR C677T polymorphism may be a risk factor for gastric cancer among Asians and Caucasians.
Assuntos
Predisposição Genética para Doença/genética , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Polimorfismo de Nucleotídeo Único/genética , Neoplasias Gástricas/genética , Povo Asiático/genética , Estudos de Casos e Controles , Humanos , Razão de Chances , Fatores de Risco , População Branca/genéticaRESUMO
Cluster of differentiation 24 (CD24) has been implicated in the development of cancer. Several single nucleotide polymorphisms (SNPs) in CD24 gene are reported to exert diverse effect on cancer risk. However, the association between CD24 SNPs and cancer risk remains unclear due to contradictory published findings. We performed a meta-analysis by pooling all available published studies on the susceptibility of CD24 rs52812045 and rs3838646 polymorphisms to cancer. The pooled odds ratios (ORs) with 95 % confidence intervals (95 % CIs) were calculated. There were five independent case-control studies with 5,539 cases and 10,241 controls included into the present study. The pooled results showed that no appreciable relationship was identified between any of the SNPs of CD24 and cancer risk. Interestingly, a protective role of the CD24 rs3838646 polymorphism was found in the risk of breast cancer, but lack of statistical significance (del allele vs. TG allele: OR = 0.89; 95 % CI, 0.79-1.01; P OR = 0.063; del/del vs. TG/TG: OR = 0.70; 95 % CI, 0.44-1.12; P OR = 0.135; del/TG vs. TG/TG: OR = 0.91; 95 % CI, 0.80-1.04, P OR = 0.180; del/del + del/TG vs. TG/TG: OR = 0.90; 95 % CI, 0.79-1.03; P OR = 0.123; del/del vs. TG/TG + del/TG: OR = 0.69; 95 % CI, 0.44-1.08, P OR = 0.105). Our study firstly provides the evidence that SNPs (rs52812045 and rs3838646) of CD24 may not modify the risk of cancer. Nonetheless, more individual studies with high quality are needed for further elucidation.
Assuntos
Antígeno CD24/genética , Predisposição Genética para Doença/genética , Neoplasias/genética , Polimorfismo de Nucleotídeo Único , Neoplasias da Mama/genética , Estudos de Casos e Controles , Feminino , Frequência do Gene , Genótipo , Humanos , Razão de Chances , Fatores de RiscoRESUMO
The circadian clock underlies daily rhythms of diverse physiological processes, and alterations in clock function have been linked to numerous pathologies. To apply chemical biology methods to modulate and dissect the clock mechanism with new chemical probes, we performed a circadian screen of â¼120,000 uncharacterized compounds on human cells containing a circadian reporter. The analysis identified a small molecule that potently lengthens the circadian period in a dose-dependent manner. Subsequent analysis showed that the compound also lengthened the period in a variety of cells from different tissues including the mouse suprachiasmatic nucleus, the central clock controlling behavioral rhythms. Based on the prominent period lengthening effect, we named the compound longdaysin. Longdaysin was amenable for chemical modification to perform affinity chromatography coupled with mass spectrometry analysis to identify target proteins. Combined with siRNA-mediated gene knockdown, we identified the protein kinases CKIδ, CKIα, and ERK2 as targets of longdaysin responsible for the observed effect on circadian period. Although individual knockdown of CKIδ, CKIα, and ERK2 had small period effects, their combinatorial knockdown dramatically lengthened the period similar to longdaysin treatment. We characterized the role of CKIα in the clock mechanism and found that CKIα-mediated phosphorylation stimulated degradation of a clock protein PER1, similar to the function of CKIδ. Longdaysin treatment inhibited PER1 degradation, providing insight into the mechanism of longdaysin-dependent period lengthening. Using larval zebrafish, we further demonstrated that longdaysin drastically lengthened circadian period in vivo. Taken together, the chemical biology approach not only revealed CKIα as a clock regulatory kinase but also identified a multiple kinase network conferring robustness to the clock. Longdaysin provides novel possibilities in manipulating clock function due to its ability to simultaneously inhibit several key components of this conserved network across species.
Assuntos
Adenina/análogos & derivados , Relógios Biológicos/efeitos dos fármacos , Proteínas CLOCK/genética , Proteínas CLOCK/metabolismo , Ritmo Circadiano/efeitos dos fármacos , Ritmo Circadiano/fisiologia , Adenina/metabolismo , Animais , Relógios Biológicos/genética , Caseína Quinase I/metabolismo , Linhagem Celular Tumoral , Ritmo Circadiano/genética , Quinases Ciclina-Dependentes/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Técnicas de Silenciamento de Genes , Estudo de Associação Genômica Ampla , Histonas/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteínas Circadianas Period/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Interferência de RNA , Fatores de Transcrição/genética , Fatores de Transcrição/fisiologia , Peixe-Zebra/genética , Peixe-Zebra/fisiologiaRESUMO
The BCR-ABL fusion gene in chromosome translocation, t (9; 22), and its product, p210BCR/ABL oncogenic tyrosine kinase, is the underlying molecular mechanism that leads to the development of CML. Quantitative detection of BCR-ABL fusion gene has become a reliable approach to diagnose and monitor CML. The aim of this study was to evaluate a Roche t (9; 22) kit in CML diagnosis, monitoring treatment responses, and identification of relapse. Using BCR-ABL fusion gene-expressing K562 cells, a series of standard samples were prepared and used to establish a curve for the calculation of BCR-ABL fusion gene expression in patient samples. Our results indicate that PCR detection system with aforementioned kit has good reproducibility. In addition, the relative concentration of BCR-ABL measured by PCR was in agreement with the patient's response to the Imatinib treatment and bone marrow morphology remission. Furthermore, we found that the relative concentration of BCR-ABL fusion gene increased 1-3 months before CML relapse was clinically and cytogenetically diagnosed, suggesting that the PCR-based BCR-ABL fusion gene detection with t (9; 22) kit is able to diagnose the recurrence of CML at least 1 month earlier than the classic cytogenetic analysis. In conclusion, detection of BCR-ABL fusion gene expression in CML using Roche t (9; 22) kit has great clinical value in the primary diagnosis, monitoring treatment responses, and identification of relapse in CML patients.
Assuntos
Proteínas de Fusão bcr-abl/genética , Proteínas de Fusão bcr-abl/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/prevenção & controle , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Antineoplásicos/uso terapêutico , Benzamidas , Biomarcadores/metabolismo , Transplante de Medula Óssea , Humanos , Mesilato de Imatinib , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Piperazinas/uso terapêutico , Pirimidinas/uso terapêutico , Kit de Reagentes para Diagnóstico , Recidiva , Indução de RemissãoRESUMO
Visceral leishmaniasis is responsible for up to 30,000 deaths every year. Current treatments have shortcomings that include toxicity and variable efficacy across endemic regions. Previously, we reported the discovery of GNF6702, a selective inhibitor of the kinetoplastid proteasome, which cleared parasites in murine models of leishmaniasis, Chagas disease, and human African trypanosomiasis. Here, we describe the discovery and characterization of LXE408, a structurally related kinetoplastid-selective proteasome inhibitor currently in Phase 1 human clinical trials. Furthermore, we present high-resolution cryo-EM structures of the Leishmania tarentolae proteasome in complex with LXE408, which provides a compelling explanation for the noncompetitive mode of binding of this novel class of inhibitors of the kinetoplastid proteasome.
Assuntos
Antiprotozoários/química , Antiprotozoários/farmacologia , Leishmaniose Visceral/tratamento farmacológico , Oxazóis/química , Oxazóis/farmacologia , Inibidores de Proteassoma/química , Inibidores de Proteassoma/farmacologia , Pirimidinas/química , Pirimidinas/farmacologia , Animais , Antiprotozoários/uso terapêutico , Cães , Humanos , Leishmania donovani/efeitos dos fármacos , Leishmania donovani/isolamento & purificação , Leishmania major/efeitos dos fármacos , Leishmania major/isolamento & purificação , Leishmaniose Visceral/parasitologia , Fígado/parasitologia , Macaca fascicularis , Camundongos , Camundongos Endogâmicos BALB C , Oxazóis/uso terapêutico , Inibidores de Proteassoma/uso terapêutico , Pirimidinas/uso terapêutico , Ratos , Ratos Sprague-Dawley , Triazóis/químicaRESUMO
Survival of Borrelia burgdorferi in ticks and mammals is facilitated, at least in part, by the selective expression of lipoproteins. Outer surface protein (Osp) A participates in spirochete adherence to the tick gut. As ospB is expressed on a bicistronic operon with ospA, we have now investigated the role of OspB by generating an OspB-deficient B. burgdorferi and examining its phenotype throughout the spirochete life cycle. Similar to wild-type isolates, the OspB-deficient B. burgdorferi were able to readily infect and persist in mice. OspB-deficient B. burgdorferi were capable of migrating to the feeding ticks but had an impaired ability to adhere to the tick gut and survive within the vector. Furthermore, the OspB-deficient B. burgdorferi bound poorly to tick gut extracts. The complementation of the OspB-deficient spirochete in trans, with a wild-type copy of ospB gene, restored its ability to bind tick gut. Taken together, these data suggest that OspB has an important role within Ixodes scapularis and that B. burgdorferi relies upon multiple genes to efficiently persist in ticks.
Assuntos
Antígenos de Bactérias/fisiologia , Aderência Bacteriana/fisiologia , Proteínas da Membrana Bacteriana Externa/fisiologia , Borrelia burgdorferi/fisiologia , Sobrevivência Celular , Ixodes/microbiologia , Animais , Antígenos de Bactérias/imunologia , Antígenos de Superfície/imunologia , Proteínas da Membrana Bacteriana Externa/imunologia , Vacinas Bacterianas/imunologia , Sistema Digestório/imunologia , Sistema Digestório/microbiologia , Imunofluorescência , Lipoproteínas/imunologia , Doença de Lyme/prevenção & controle , Doença de Lyme/transmissão , Camundongos , Camundongos Endogâmicos C3H , Mutação , Reação em Cadeia da Polimerase , Spirochaetales/imunologia , Spirochaetales/metabolismoRESUMO
Macrophage-orchestrated, low-grade chronic inflammation plays a pivotal role in obesity and atherogenesis. However, the underlying regulatory mechanisms remain incompletely understood. Here, we identify major vault protein (MVP), the main component of unique cellular ribonucleoprotein particles, as a suppressor for NF-κB signaling in macrophages. Both global and myeloid-specific MVP gene knockout aggravates high-fat diet induced obesity, insulin resistance, hepatic steatosis and atherosclerosis in mice. The exacerbated metabolic disorders caused by MVP deficiency are accompanied with increased macrophage infiltration and heightened inflammatory responses in the microenvironments. In vitro studies reveal that MVP interacts with TRAF6 preventing its recruitment to IRAK1 and subsequent oligomerization and ubiquitination. Overexpression of MVP and its α-helical domain inhibits the activity of TRAF6 and suppresses macrophage inflammation. Our results demonstrate that macrophage MVP constitutes a key constraint of NF-κB signaling thereby suppressing metabolic diseases.
Assuntos
Aterosclerose/imunologia , Fígado Gorduroso/imunologia , Inflamação/imunologia , Macrófagos/imunologia , Obesidade/imunologia , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Tecido Adiposo/patologia , Animais , Aterosclerose/etiologia , Aterosclerose/metabolismo , Biópsia , Células da Medula Óssea , Dieta Hiperlipídica/efeitos adversos , Modelos Animais de Doenças , Fígado Gorduroso/etiologia , Fígado Gorduroso/metabolismo , Feminino , Técnicas de Inativação de Genes , Humanos , Quinase I-kappa B/metabolismo , Inflamação/etiologia , Quinases Associadas a Receptores de Interleucina-1/metabolismo , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , NF-kappa B/metabolismo , Obesidade/etiologia , Obesidade/metabolismo , Obesidade/patologia , Cultura Primária de Células , Transdução de Sinais/imunologia , Fator 6 Associado a Receptor de TNF/metabolismo , Ubiquitinação , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética , Partículas de Ribonucleoproteínas em Forma de Abóbada/imunologiaRESUMO
32P high-dose rate brachytherapy allows high-dose radiation delivery to target lesions with less damage to adjacent tissues. The early evaluation of its therapeutic effect on tumours is vital for the optimization of treatment regimes. The most commonly used 32P-CP colloid tends to leak with blind therapeutic area after intratumour injection. We prepared 32P-chromic phosphate-polylactide-co-glycolide (32P-CP-PLGA) seeds with biodegradable PLGA as a framework and investigated their characteristics in vitro and in vivo. We also evaluated the therapeutic effect of 32P-CP-PLGA brachytherapy for glioma with the integrin αvß3-targeted radiotracer 68Ga-3PRGD2. 32P-CP-PLGA seeds (seed group, SG, 185 MBq) and 32P-CP colloid (colloid group, CG, 18.5 MBq) were implanted or injected into human glioma xenografts in nude mice. Scanning electron microscopy (SEM) of the seeds, micro-SPECT imaging, and biodistribution studies were performed at different time points. The tumour volume was measured using a caliper, and 68Ga-3PRGD2 micro-PET-CT imaging was performed to evaluate the therapeutic effect after 32P intratumour administration. The delayed release of 32P-CP was observed with biodegradation of vehicle PLGA. Intratumoural effective half-life of 32P-CP in the SG (13.3 ± 0.3) d was longer than that in the CG (10.4 ± 0.3) d (P < 0.05), with liver appearance in the CG on SPECT. A radioactivity gradient developed inside the tumour in the SG, as confirmed by micro-SPECT and SEM. Tumour uptake of 68Ga-3PRGD2 displayed a significant increase on day 0.5 in the SG and decreased earlier (on day 2) than the volume reduction (on day 8). Thus, 32P-CP-PLGA seeds, controlling the release of entrapped 32P-CP particles, are promising for glioma brachytherapy, and 68Ga-3PRGD2 imaging shows potential for early response evaluation of 32P-CP-PLGA seeds brachytherapy.
Assuntos
Braquiterapia/métodos , Glioma/terapia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/farmacocinética , Compostos Radiofarmacêuticos/química , Animais , Compostos de Cromo , Meia-Vida , Xenoenxertos , Humanos , Camundongos , Microscopia Eletrônica de Varredura , Fosfatos , Radioisótopos de Fósforo , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Compostos Radiofarmacêuticos/farmacocinética , Distribuição Tecidual , Tomografia Computadorizada de Emissão de Fóton ÚnicoRESUMO
OBJECTIVE: Graft size is one of the major risk factors in adult-to-adult living donor liver transplantation and rapid regeneration is an essential post-operative requirement. Ischemic preconditioning (IPC) has been shown to be an effective strategy in the reduction of hepatic ischemia-reperfusion injury and stimulation of liver regeneration. This study was designed to evaluate the effects of IPC on liver regeneration in small-for-size liver grafts. METHODS: We employed a rat orthotopic liver transplantation model using small-for-size (30%) grafts, in the presence or absence (control) of IPC (10 min of ischemia followed by 15 min of reperfusion). Survival rate, graft injury, hepatocellular proliferation, cell cycle progression, Stat3 activation, as well as TNF-alpha and IL-6 expression were assessed. RESULTS: IPC significantly enhanced the extent of graft injury and hindered hepatic regeneration in small-for-size liver grafts. The 7-day survival rate was also reduced by IPC, but failed to reach statistical significance. IPC did not affect TNF-alpha levels, but significantly decreased the elevation of IL-6 after reperfusion. These findings were correlated with down-regulation of cyclin E and cyclin D1, and decreased numbers of PCNA-positive nuclei in IPC grafts. These results were inconsistent with Stat3 activation, as P-Stat3 exhibited a stronger and prolonged pattern of expression in the IPC group, compared to controls. CONCLUSIONS: Ischemic preconditioning may impair liver regeneration in small-for-size liver grafts by decreasing IL-6 and blunting cell cycle progression, through a mechanism at least partially independent of Stat3.
Assuntos
Precondicionamento Isquêmico , Regeneração Hepática , Transplante de Fígado , Animais , Ciclo Celular , Interleucina-6/fisiologia , Fígado/patologia , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Modelos Animais , Antígeno Nuclear de Célula em Proliferação/análise , Ratos , Ratos Endogâmicos Lew , Fator de Transcrição STAT3/metabolismo , Taxa de Sobrevida , Fator de Necrose Tumoral alfa/fisiologiaRESUMO
Small-airway remodelling is one of the most remarkable pathological features of chronic obstructive pulmonary disease (COPD), in which angiogenesis plays a critical role that contributes to disease progression. The endothelial cell-specific mitogen vascular endothelial growth factor (VEGF), as well as its receptors, VEGFR1, VEGFR2, are thought to be the major mediators of pathological angiogenesis, and sunitinib exhibits anti-angiogenesis property through VEGF blockage and has been widely used to treat various cancers. In our study, Sprague-Dawley rats were subjected to lipopolysaccharide (LPS) injection and cigarette smoke (CS) inhalation to induce COPD, following sunitinib administration was conducted. Haematoxylin-eosin, Masson staining and immunostaining analysis were used to evaluate the pathological changes; quantitative real-time PCR and enzyme-linked immunosorbent assay were performed to provide more compelling data on the function of VEGF, VEGFR1, VEGFR2 in angiogenesis. Sunitinib treatment was associated with less angiogenesis in small-airway remodelling with a slightly disordered lung architecture, and lower expression level of VEGF, VEGFR1, VEGFR2. Overall, our results indicate that VEGF is a vital important factor that contributes to the small-airway remodelling in a rat model of COPD through promoting angiogenesis, which mainly depend on the specific binding between VEGF and VEGFR1 and can be effectively attenuated by sunitinib.
Assuntos
Remodelação das Vias Aéreas , Inibidores da Angiogênese/administração & dosagem , Indóis/administração & dosagem , Neovascularização Patológica/metabolismo , Doença Pulmonar Obstrutiva Crônica/metabolismo , Pirróis/administração & dosagem , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Modelos Animais de Doenças , Lipopolissacarídeos/administração & dosagem , Masculino , Doença Pulmonar Obstrutiva Crônica/induzido quimicamente , Doença Pulmonar Obstrutiva Crônica/prevenção & controle , Ratos Sprague-Dawley , Fumaça , Sunitinibe , Nicotiana/toxicidade , Receptor 1 de Fatores de Crescimento do Endotélio Vascular/metabolismo , Receptor 2 de Fatores de Crescimento do Endotélio Vascular/metabolismoRESUMO
Borrelia burgdorferi survives in an enzootic cycle, and Dps proteins protect DNA against damage during starvation or oxidative stress. The role of a Dps homologue encoded by Borrelia in spirochaete survival was assessed. Dps-deficient spirochaetes were infectious in mice via needle-inoculation at the dose of 10(5) spirochaetes. Larval ticks successfully acquired Dps-deficient spirochaetes via a blood meal on mice. However, after extended periods within unfed nymphs, the Dps-deficient spirochaetes failed to be transmitted to a new host when nymphs fed. Our data suggest that Dps functions to protect the spirochaetes during dormancy in unfed ticks, and in its absence, the spirochaetes become susceptible during tick feeding. dps is differentially expressed in vivo- low in mice and high in ticks - but constitutively expressed in vitro, showing little change during growth or in response to oxidative stress. Borrelia Dps forms a dodecameric complex capable of sequestering iron. The Dps-deficient spirochaetes showed no defect in starvation and oxidative stress assays, perhaps due to the lack of iron in spirochaetes grown in vitro. Dps is critical for spirochaete persistence within ticks, and strategies to interfere with Dps could potentially reduce Borrelia populations in nature and thereby influence the incidence of Lyme disease.
Assuntos
Proteínas de Bactérias/metabolismo , Borrelia burgdorferi/fisiologia , Proteínas de Ligação a DNA/metabolismo , Ixodes/microbiologia , Doença de Lyme/microbiologia , Sequência de Aminoácidos , Animais , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Teste de Complementação Genética , Ixodes/crescimento & desenvolvimento , Doença de Lyme/transmissão , Camundongos , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Ninfa/microbiologia , Fenótipo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Transformação GenéticaRESUMO
Borrelia burgdorferi preferentially induces selected genes in mice or ticks, and studies suggest that ospD is down-regulated in response to host-specific signals. We now directly show that ospD expression is generally elevated within Ixodes scapularis compared with mice. We then assessed the importance of OspD throughout the spirochete life cycle by generating OspD-deficient B. burgdorferi and examining the mutant in the murine model of tick-transmitted Lyme borreliosis. The lack of OspD did not influence B. burgdorferi infectivity in mice or the acquisition of spirochetes by I. scapularis. OspD adhered to tick gut extracts in vitro, and the OspD-deficient B. burgdorferi strain had a threefold decrease in colonization of the tick gut in vivo. This decrease, however, did not alter subsequent spirochete transmission during a second blood meal. These data suggest that B. burgdorferi can compensate for the lack of OspD in both ticks and mice and that OspD may have a nonessential, secondary, role in B. burgdorferi persistence within I. scapularis.
Assuntos
Proteínas da Membrana Bacteriana Externa/fisiologia , Borrelia burgdorferi/crescimento & desenvolvimento , Borrelia burgdorferi/fisiologia , Ixodes/microbiologia , Doença de Lyme/microbiologia , Animais , Proteínas da Membrana Bacteriana Externa/biossíntese , Proteínas da Membrana Bacteriana Externa/genética , Borrelia burgdorferi/genética , Modelos Animais de Doenças , Feminino , Regulação Bacteriana da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos C3H , MutaçãoRESUMO
BBK32, a fibronectin-binding protein of Borrelia burgdorferi, is one of many surface lipoproteins that are differentially expressed by the Lyme disease spirochete at various stages of its life cycle. The level of BBK32 expression in B. burgdorferi is highest during infection of the mammalian host and lowest in flat ticks. This temporal expression profile, along with its fibronectin-binding activity, strongly suggests that BBK32 may play an important role in Lyme pathogenesis in the host. To test this hypothesis, we constructed an isogenic BBK32 deletion mutant from wild-type B. burgdorferi B31 by replacing the BBK32 gene with a kanamycin resistance cassette through homologous recombination. We examined both the wild-type strain and the BBK32 deletion mutant extensively in the experimental mouse-tick model of the Borrelia life cycle. Our data indicated that B. burgdorferi lacking BBK32 retained full pathogenicity in mice, regardless of whether mice were infected artificially by syringe inoculation or naturally by tick bite. The loss of BBK32 expression in the mutant had no adverse effect on spirochete acquisition (mouse-to-tick) and transmission (tick-to-mouse) processes. These results suggest that additional B. burgdorferi proteins can complement the function of BBK32, fibronectin binding or otherwise, during the natural spirochete life cycle.