RESUMO
OBJECTIVES: Penile squamous cell carcinomas (PCs) are rare malignancies with a dismal prognosis in a metastatic setting; therefore, novel immunotherapeutic modalities are an unmet need. One such modality is the immune checkpoint molecule programmed cell death ligand 1 (PD-L1). We sought to analyze PD-L1 expression and its correlation with various clinicopathologic parameters in a contemporary cohort of 134 patients with PC. METHODS: A cohort of 134 patients with PC was studied for PD-L1 immunohistochemistry. The PD-L1 expression was evaluated using a combined proportion score with a cutoff of 1 or higher to define positivity. The results were correlated with various clinicopathologic parameters. RESULTS: Overall, 77 (57%) patients had positive PD-L1 expression. Significantly high PD-L1 expression was observed in high-grade tumors (P = .006). We found that 37% of human papillomavirus (HPV)-associated subtypes and 73% of other histotype tumors expressed PD-L1, while 63% of HPV-associated tumors and 27% of other histotype tumors did not (odds ratio, 1.35; P = .002 when compared for HPV-associated groups vs all others). Similarly, PD-L1-positive tumors had a 3.61-times higher chance of being node positive than PD-L1-negative tumors (P = .0009). In addition, PD-L1 high-positive tumors had a 5-times higher chance of being p16ink4a negative than PD-L1 low-positive tumors (P = .004). The PD-L1-positive tumors had a lower overall survival and cancer-specific survival than PD-L1-negative tumors. CONCLUSIONS: Overall, PD-L1 expression is associated with high-grade and metastatic tumors. Lower PD-L1 expression is observed more frequently in HPV-associated (warty or basaloid) subtypes than in other, predominantly HPV-independent types. As a result, PD-L1 positivity, including higher expression, portends lower overall and cancer-specific survival. These data provide a rational for further investigating PD-L1-based immunotherapeutics in PC.
Assuntos
Carcinoma de Células Escamosas , Infecções por Papillomavirus , Neoplasias Penianas , Masculino , Humanos , Infecções por Papillomavirus/complicações , Infecções por Papillomavirus/metabolismo , Antígeno B7-H1/metabolismo , Ligantes , Prognóstico , Carcinoma de Células Escamosas/patologia , Neoplasias Penianas/patologia , Apoptose , Biomarcadores Tumorais/metabolismoRESUMO
Abdominal aortic aneurysm (AAA) formation is characterized by inflammation, leukocyte infiltration, and vascular remodeling. Resolvin D1 (RvD1) is derived from ω-3 polyunsaturated fatty acids and is involved in the resolution phase of chronic inflammatory diseases. The aim of this study was to decipher the protective role of RvD1 via formyl peptide receptor 2 (FPR2) receptor signaling in attenuating abdominal aortic aneurysms (AAA). The elastase-treatment model of AAA in C57BL/6 (WT) mice and human AAA tissue was used to confirm our hypotheses. Elastase-treated FPR2-/- mice had a significant increase in aortic diameter, proinflammatory cytokine production, immune cell infiltration (macrophages and neutrophils), elastic fiber disruption, and decrease in smooth muscle cell α-actin expression compared to elastase-treated WT mice. RvD1 treatment attenuated AAA formation, aortic inflammation, and vascular remodeling in WT mice, but not in FPR2-/- mice. Importantly, human AAA tissue demonstrated significantly decreased FPR2 mRNA expression compared to non-aneurysm human aortas. Mechanistically, RvD1/FPR2 signaling mitigated p47phox phosphorylation and prevented hallmarks of ferroptosis, such as lipid peroxidation and Nrf2 translocation, thereby attenuating HMGB1 secretion. Collectively, this study demonstrates RvD1-mediated immunomodulation of FPR2 signaling on macrophages to mitigate ferroptosis and HMGB1 release, leading to resolution of aortic inflammation and remodeling during AAA pathogenesis.
Assuntos
Aneurisma da Aorta Abdominal , Ferroptose , Proteína HMGB1 , Actinas/metabolismo , Animais , Aneurisma da Aorta Abdominal/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Ácidos Docosa-Hexaenoicos/metabolismo , Proteína HMGB1/metabolismo , Humanos , Inflamação/metabolismo , Macrófagos/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fator 2 Relacionado a NF-E2/metabolismo , Elastase Pancreática/metabolismo , RNA Mensageiro/metabolismo , Receptores de Formil Peptídeo/genética , Receptores de Formil Peptídeo/metabolismo , Receptores de Lipoxinas , Remodelação VascularRESUMO
Aerobic Escherichia coli growth at restricted iron concentrations (≤ 1.75 ± 0.04 µM) is characterized by lower biomass yield, higher acetate accumulation and higher activation of the siderophore iron-acquisition systems. Although iron homeostasis in E. coli has been studied intensively, previous studies focused only on understanding the regulation of the iron import systems and the iron-requiring enzymes. Here, the effect of iron availability on the energy metabolism of E. coli has been investigated. It was established that aerobic cultures growing under limiting iron conditions showed lower ATP yield per glucose, lower growth rate and lower TCA cycle activity and respiration, at the same time as increased glucose consumption, acetate and pyruvate accumulation, practically mimicking microaerobic growth. However, at excess iron, independent of oxygen availability, the cultures showed high cellular energetics (5.8 ATP/mol of glucose) by using pathways requiring iron-rich complex proteins found in the TCA cycle and respiratory chain. In conditions of iron excess, some iron-requiring terminal reductases of the respiratory chain, that were thought to function only under anaerobiosis, were used by the E. coli, when in aerobic conditions, to maintain high respiratory activity. This allowed it to produce more biomass and more reactive oxygen species that were controlled by the higher activity of the antioxidant defenses (SOD, peroxidase and catalase) and the iron-sulfur cluster repair systems.
Assuntos
Proteínas de Escherichia coli , Escherichia coli , Trifosfato de Adenosina , Anaerobiose , Transporte de Elétrons , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Regulação Bacteriana da Expressão Gênica , Glucose/metabolismo , Ferro/metabolismo , Oxirredutases/metabolismoRESUMO
Pannexin-1 (Panx1) channels have been shown to regulate leukocyte trafficking and tissue inflammation but the mechanism of Panx1 in chronic vascular diseases like abdominal aortic aneurysms (AAA) is unknown. Here we demonstrate that Panx1 on endothelial cells, but not smooth muscle cells, orchestrate a cascade of signaling events to mediate vascular inflammation and remodeling. Mechanistically, Panx1 on endothelial cells acts as a conduit for ATP release that stimulates macrophage activation via P2X7 receptors and mitochondrial DNA release to increase IL-1ß and HMGB1 secretion. Secondly, Panx1 signaling regulates smooth muscle cell-dependent intracellular Ca2+ release and vascular remodeling via P2Y2 receptors. Panx1 blockade using probenecid markedly inhibits leukocyte transmigration, aortic inflammation and remodeling to mitigate AAA formation. Panx1 expression is upregulated in human AAAs and retrospective clinical data demonstrated reduced mortality in aortic aneurysm patients treated with Panx1 inhibitors. Collectively, these data identify Panx1 signaling as a contributory mechanism of AAA formation.
Assuntos
Aneurisma da Aorta Abdominal , Células Endoteliais , Trifosfato de Adenosina/metabolismo , Aneurisma da Aorta Abdominal/genética , Conexinas/genética , Conexinas/metabolismo , Células Endoteliais/metabolismo , Humanos , Inflamação/metabolismo , Macrófagos/metabolismo , Miócitos de Músculo Liso/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Estudos RetrospectivosRESUMO
BACKGROUND: Thoracic aortic aneurysms (TAA) are a progressive disease characterized by inflammation, smooth muscle cell activation and matrix degradation. We hypothesized that mesenchymal stem cells (MSCs) can immunomodulate vascular inflammation and remodeling via altered microRNA (miRNAs) expression profile to attenuate TAA formation. MATERIALS AND METHODS: C57BL/6 mice underwent topical elastase application to form descending TAAs. Mice were also treated with MSCs on days 1 and 5 and aortas were analyzed on day 14 for aortic diameter. Cytokine array was performed in aortic tissue and total RNA was tagged and hybridized for miRNAs microarray analysis. Immunohistochemistry was performed for elastin degradation and leukocyte infiltration. RESULTS: Treatment with MSCs significantly attenuated aortic diameter and TAA formation compared to untreated mice. MSC administration also attenuated T-cell, neutrophil and macrophage infiltration and prevented elastic degradation to mitigate vascular remodeling. MSC treatment also attenuated aortic inflammation by decreasing proinflammatory cytokines (CXCL13, IL-27, CXCL12 and RANTES) and upregulating anti-inflammatory interleukin-10 expression in aortic tissue of elastase-treated mice. TAA formation demonstrated activation of specific miRNAs that are associated with aortic inflammation and vascular remodeling. Our results also demonstrated that MSCs modulate a different set of miRNAs that are associated with decrease leukocyte infiltration and vascular inflammation to attenuate the aortic diameter and TAA formation. CONCLUSIONS: These results indicate that MSCs immunomodulate specific miRNAs that are associated with modulating hallmarks of aortic inflammation and vascular remodeling of aortic aneurysms. Targeted therapies designed using MSCs and miRNAs have the potential to regulate the growth and development of TAAs.
Assuntos
Aneurisma da Aorta Torácica , Células-Tronco Mesenquimais , MicroRNAs , Animais , Aneurisma da Aorta Torácica/terapia , Modelos Animais de Doenças , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Asthma is a common inflammatory lung disease with no known cure. Previously, we uncovered a lung TNFR2+ conventional DC2 subset (cDC2s) that induces regulatory T cells (Tregs) maintaining lung tolerance at steady state but promotes TH2 response during house dust mite (HDM)-induced asthma. Lung IFNß is essential for TNFR2+ cDC2s-mediated lung tolerance. Here, we showed that exogenous IFNß reprogrammed TH2-promoting pathogenic TNFR2+ cDC2s back to tolerogenic DCs, alleviating eosinophilic asthma and preventing asthma exacerbation. Mechanistically, inhaled IFNß, not IFNα, activated ERK2 signaling in pathogenic lung TNFR2+ cDC2s, leading to enhanced fatty acid oxidation (FAO) and lung Treg induction. Last, human IFNß reprogrammed pathogenic human lung TNFR2+ cDC2s from patients with emphysema ex vivo. Thus, we identified an IFNß-specific ERK2-FAO pathway that might be harnessed for DC therapy.
Assuntos
Asma/imunologia , Células Dendríticas/transplante , Interferon beta/uso terapêutico , Receptores Tipo II do Fator de Necrose Tumoral/genética , Células Th2/imunologia , Transferência Adotiva , Animais , Asma/patologia , Células Cultivadas , Células Dendríticas/imunologia , Dermatophagoides pteronyssinus/imunologia , Modelos Animais de Doenças , Ácidos Graxos/metabolismo , Humanos , Tolerância Imunológica/imunologia , Fatores Imunológicos/uso terapêutico , Interferon-alfa/farmacologia , Pulmão/citologia , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Oxirredução , Receptor de Interferon alfa e beta/genética , Transdução de Sinais/imunologia , Linfócitos T Reguladores/imunologiaRESUMO
Cell lines used in bioproduction are routinely engineered to improve their production efficiency. Numerous strategies, such as random mutagenesis, RNA interference screens, and transcriptome analyses have been employed to identify effective engineering targets. A genome-wide small interfering RNA screen previously identified the CASP8AP2 gene as a potential engineering target for improved expression of recombinant protein in the HEK293 cell line. Here, we validate the CASP8AP2 gene as an engineering target in HEK293 cells by knocking it out using CRISPR/Cas9 genome editing and assessing the effect of its knockout on recombinant protein expression, cell growth, cell viability, and overall gene expression. HEK293 cells lacking CASP8AP2 showed a seven-fold increase in specific expression of recombinant luciferase and a 2.5-fold increase in specific expression of recombinant SEAP, without significantly affecting cell growth and viability. Transcriptome analysis revealed that the deregulation of the cell cycle, specifically the upregulation of the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene, contributed to the improvement in recombinant protein expression in CASP8AP2 deficient cells. The results validate the CASP8AP2 gene is a viable engineering target for improved recombinant protein expression in the HEK293 cell line.
Assuntos
Proteínas Reguladoras de Apoptose/deficiência , Proteínas de Ligação ao Cálcio/deficiência , Inibidor p16 de Quinase Dependente de Ciclina/biossíntese , Regulação Enzimológica da Expressão Gênica , Regulação para Cima , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genéticaRESUMO
BACKGROUND: Our hypothesis is that the immunomodulatory capacities of mesenchymal stem cellâderived extracellular vesicles (EVs) can be enhanced by specific microRNAs (miRNAs) to effectively attenuate post-transplant lung ischemiaâreperfusion (IR) injury. METHODS: The expression of miR-206 was analyzed in bronchoalveolar lavage (BAL) fluid of patients on Days 0 and 1 after lung transplantation. Lung IR injury was evaluated in C57BL/6 mice using a left lung hilar-ligation model with or without treatment with EVs or antagomiR-206âenriched EVs. Murine lung tissue was used for miRNA microarray hybridization analysis, and cytokine expression, lung injury, and edema were evaluated. A donation after circulatory death and murine orthotopic lung transplantation model was used to evaluate the protection by enriched EVs against lung IR injury. In vitro studies analyzed type II epithelial cell activation after coculturing with EVs. RESULTS: A significant upregulation of miR-206 was observed in the BAL fluid of patients on Day 1 after lung transplantation compared with Day 0 and in murine lungs after IR injury compared with sham. Treatment with antagomiR-206âenriched EVs attenuated lung dysfunction, injury, and edema compared with treatment with EVs alone after murine lung IR injury. Enriched EVs reduced lung injury and neutrophil infiltration as well as improved allograft oxygenation after murine orthotopic lung transplantation. Enriched EVs significantly decreased proinflammatory cytokines, especially epithelial cellâdependent CXCL1 expression, in the in vivo and in vitro IR injury models. CONCLUSIONS: EVs can be used as biomimetic nanovehicles for protective immunomodulation by enriching them with antagomiR-206 to mitigate epithelial cell activation and neutrophil infiltration in the lungs after IR injury.
Assuntos
Antagomirs/genética , Quimiocina CXCL1/genética , Regulação da Expressão Gênica , Lesão Pulmonar/prevenção & controle , MicroRNAs/genética , Traumatismo por Reperfusão/prevenção & controle , Animais , Antagomirs/biossíntese , Líquido da Lavagem Broncoalveolar , Quimiocina CXCL1/biossíntese , Modelos Animais de Doenças , Humanos , Lesão Pulmonar/genética , Lesão Pulmonar/metabolismo , Transplante de Pulmão , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/biossíntese , RNA/genética , Traumatismo por Reperfusão/genética , Traumatismo por Reperfusão/metabolismoRESUMO
Abdominal aortic aneurysm (AAA) formation is characterized by inflammation, leukocyte infiltration, and vascular remodeling. This study investigates the role of TRPV4 channels, which are transmembrane calcium channels that can regulate vascular tone, in modulating AAA formation. The elastase-treatment model of AAA in C57BL6 (WT) mice and Angiotensin II treatment model in ApoE-/- mice were used to confirm our hypotheses. The administration of a specific TRPV4 antagonist, GSK2193874, in elastase-treated WT mice and in AngII-treated ApoE-/- mice caused a significant attenuation of aortic diameter, decrease in pro-inflammatory cytokines (IL-1ß, IL-6, IL-17, MCP-1, MIP-1α, MIP-2, RANTES, and TNF-α), inflammatory cell infiltration (CD3 + T cells, macrophages, and neutrophils), elastic fiber disruption, and an increase in smooth muscle cell α-actin expression compared to untreated mice. Similarly, elastase-treated TRPV4-/- mice had a significant decrease in AAA formation, aortic inflammation, and vascular remodeling compared to elastase-treated WT mice on Day 14. In vitro studies demonstrated that the inhibition of TRPV4 channels mitigates aortic smooth muscle cell-dependent inflammatory cytokine production as well as decreases neutrophil transmigration through aortic endothelial cells. Therefore, our results suggest that TRPV4 antagonism can attenuate aortic inflammation and remodeling via decreased smooth muscle cell activation and neutrophil transendothelial migration during AAA formation.
Assuntos
Aneurisma da Aorta Abdominal/prevenção & controle , Inflamação/prevenção & controle , Macrófagos/efeitos dos fármacos , Piperidinas/farmacologia , Quinolinas/farmacologia , Canais de Cátion TRPV/antagonistas & inibidores , Animais , Aneurisma da Aorta Abdominal/etiologia , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/patologia , Inflamação/etiologia , Inflamação/metabolismo , Inflamação/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Elastase Pancreática/metabolismoRESUMO
OBJECTIVE: We tested the hypothesis that systemic administration of an A2AR agonist will reduce multiorgan IRI in a porcine model of ECPR. SUMMARY BACKGROUND DATA: Advances in ECPR have decreased mortality after cardiac arrest; however, subsequent IRI contributes to late multisystem organ failure. Attenuation of IRI has been reported with the use of an A2AR agonist. METHODS: Adult swine underwent 20 minutes of circulatory arrest, induced by ventricular fibrillation, followed by 6 hours of reperfusion with ECPR. Animals were randomized to vehicle control, low-dose A2AR agonist, or high-dose A2AR agonist. A perfusion specialist using a goal-directed resuscitation protocol managed all the animals during the reperfusion period. Hourly blood, urine, and tissue samples were collected. Biochemical and microarray analyses were performed to identify differential inflammatory markers and gene expression between groups. RESULTS: Both the treatment groups demonstrated significantly higher percent reduction from peak lactate after reperfusion compared with vehicle controls. Control animals required significantly more fluid, epinephrine, and higher final pump flow while having lower urine output than both the treatment groups. The treatment groups had lower urine NGAL, an early marker of kidney injury (P = 0.01), lower plasma aspartate aminotransferase, and reduced rate of troponin rise (P = 0.01). Pro-inflammatory cytokines were lower while anti-inflammatory cytokines were significantly higher in the treatment groups. CONCLUSIONS: Using a novel and clinically relevant porcine model of circulatory arrest and ECPR, we demonstrated that a selective A2AR agonist significantly attenuated systemic IRI and warrants clinical investigation.
Assuntos
Agonistas do Receptor A2 de Adenosina/uso terapêutico , Reanimação Cardiopulmonar/efeitos adversos , Parada Cardíaca/terapia , Traumatismo por Reperfusão/prevenção & controle , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Feminino , Parada Cardíaca/complicações , Masculino , Traumatismo por Reperfusão/etiologia , SuínosRESUMO
BACKGROUND: Today's declining federal budget for scientific research is making it consistently more difficult to become federally funded. We hypothesized that even in this difficult era, surgeon-scientists have remained among the most productive and impactful researchers in lung transplantation. METHODS: Grants awarded by the NIH for the study of lung transplantation between 1985 and 2015 were identified by searching NIH RePORTER for 5 lung transplantation research areas. A grant impact metric was calculated for each grant by dividing the sum of impact factors for all associated manuscripts by the total funding for that grant. We used nonparametric univariate analysis to compare grant impact metrics by department. RESULTS: We identified 109 lung transplantation grants, totaling approximately $300 million, resulting in 2304 papers published in 421 different journals. Surgery has the third highest median grant impact metric (4.2 per $100,000). The department of surgery had a higher median grant impact metric compared to private companies (P <.0001). There was no statistical difference in the grant impact metric compared to all other medical specialties, individual departments with multiple grants, or all basic science departments (all P >.05). CONCLUSIONS: Surgeon-scientists in the field of lung transplantation have received fewer grants and less total funding compared to other researchers but have maintained an equally high level of productivity and impact. The dual-threat academic surgeon-scientist is an important asset to the research community and should continue to be supported by the NIH.
Assuntos
Pesquisa Biomédica/organização & administração , Administração Financeira/métodos , Organização do Financiamento , Transplante de Pulmão , Cirurgiões , Humanos , Estudos Retrospectivos , Estados UnidosRESUMO
Objective- The goal of this study was to determine the role of ZFP148 (zinc-finger protein 148) in aneurysm formation. Approach and Results- ZFP148 mRNA expression increased at day 3, 7, 14, 21, and 28 after during abdominal aortic aneurysm formation in C57BL/6 mice. Loss of ZFP148 conferred abdominal aortic aneurysm protection using ERTCre+ ZFP148 flx/flx mice. In a third set of experiments, smooth muscle-specific loss of ZFP148 alleles resulted in progressively greater protection using novel transgenic mice (MYH [myosin heavy chain 11] Cre+ flx/flx, flx/wt, and wt/wt). Elastin degradation, LGAL3, and neutrophil staining were significantly attenuated, while α-actin staining was increased in ZFP148 knockout mice. Results were verified in total cell ZFP148 and smooth muscle-specific knockout mice using an angiotensin II model. ZFP148 smooth muscle-specific conditional mice demonstrated increased proliferation and ZFP148 was shown to bind to the p21 promoter during abdominal aortic aneurysm formation. ZFP148 smooth muscle-specific conditional knockout mice also demonstrated decreased apoptosis as measured by decreased cleaved caspase-3 staining. ZFP148 bound smooth muscle marker genes via chromatin immunoprecipitation analysis mediated by NF-1 (neurofibromin 1) promote histone H3K4 deacetylation via histone deacetylase 5. Transient transfections and chromatin immunoprecipitation analyses demonstrated that NF-1 was required for ZFP148 protein binding to smooth muscle marker genes promoters during aneurysm formation. Elimination of NF-1 using shRNA approaches demonstrated that NF-1 is required for binding and elimination of NF-1 increased BRG1 recruitment, the ATPase subunit of the SWI/SWF complex, and increased histone acetylation. Conclusions- ZFP148 plays a critical role in multiple murine models of aneurysm formation. These results suggest that ZFP148 is important in the regulation of proliferation, smooth muscle gene downregulation, and apoptosis in aneurysm development.
Assuntos
Aneurisma da Aorta Abdominal/etiologia , Proteínas de Ligação a DNA/metabolismo , Miócitos de Músculo Liso/metabolismo , Neurofibromina 1/metabolismo , Fatores de Transcrição/metabolismo , Acetilação , Angiotensina II/farmacologia , Animais , Aneurisma da Aorta Abdominal/metabolismo , Apoptose , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/genética , Feminino , Histonas/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Killer-Antagonista Homóloga a bcl-2/genéticaRESUMO
OBJECTIVE: Resolvins have been shown to attenuate inflammation, whereas NETosis, the process of neutrophils releasing neutrophil extracellular traps (NETs), produces increased inflammation. It is hypothesized that treatment of animals with resolvin D1 (RvD1) would reduce abdominal aortic aneurysm (AAA) formation by inhibiting NETosis. METHODS: Wild-type 8- to 12-week-old C57BL/6 male mice (n = 47) and apolipoprotein E-deficient (ApoE-/-) mice (n = 20) were used in two models to demonstrate the effects of RvD1 on AAA growth. In the topical elastase AAA model, wild-type mice were divided into three groups: a deactivated elastase control group, in which sham surgery was performed using deactivated elastase and mice were intravenously injected with phosphate-buffered saline (PBS) once a day until harvest; an elastase group, in which active elastase was used to induce AAA and mice were injected with PBS daily until harvest; and an RvD1-treated group, in which AAA was induced and mice were injected with RvD1 daily until harvest. In the angiotensin II (Ang II)-induced AAA model, ApoE-/- mice were fed a high-fat diet and implanted with osmotic infusion pumps containing Ang II (1000 ng/kg/min). The Ang II model was divided into two groups: an Ang II control group, in which Ang II was delivered and mice were injected with PBS daily until harvest; and an RvD1-treated group, in which Ang II was delivered and mice were injected with RvD1 daily until harvest. On postoperative day 3, day 14, or day 28, aortic and blood samples were collected for Western blot, histology, cytokine array, enzyme-linked immunosorbent assay, and gelatin zymography after aortic diameter measurement. RESULTS: The day 14 RvD1-treated group demonstrated 42% reduced AAA diameter compared with the elastase group (P < .001). On postoperative day 3, the RvD1-treated group showed decreased levels of NETosis markers citrullinated histone H3 (P = .04) and neutrophil elastase (P = .002) compared with the elastase group. Among important cytokines involved in AAA formation, interleukin (IL) 1ß was downregulated (P = .02) whereas IL-10, a protective cytokine, was upregulated (P = .01) in the RvD1-treated group. Active matrix metalloproteinase 2 also decreased in the RvD1-treated group (P = .03). The RvD1-treated group in the Ang II AAA model, a second model, demonstrated reduced AAA diameter compared with the Ang II control group on day 28 (P < .046). The RvD1-treated group showed decreased levels of citrullinated histone H3 on day 3 (P = .002). Cytokines interferon γ, IL-1ß, C-X-C motif chemokine ligand 10, monocyte chemotactic protein 1, and regulated on activation, normal T cell expressed and secreted (RANTES) were all decreased on day 28 (P < .05). CONCLUSIONS: RvD1-mediated inhibition of NETosis may represent a future medical treatment for the attenuation of AAA growth.
Assuntos
Anti-Inflamatórios/farmacologia , Aorta Abdominal/efeitos dos fármacos , Aneurisma da Aorta Abdominal/prevenção & controle , Ácidos Docosa-Hexaenoicos/farmacologia , Armadilhas Extracelulares/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Animais , Aorta Abdominal/metabolismo , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/metabolismo , Aneurisma da Aorta Abdominal/patologia , Citrulinação , Citocinas/metabolismo , Modelos Animais de Doenças , Armadilhas Extracelulares/metabolismo , Histonas/metabolismo , Mediadores da Inflamação/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Neutrófilos/metabolismo , Neutrófilos/patologia , Elastase Pancreática , Remodelação Vascular/efeitos dos fármacosRESUMO
BACKGROUND: Tamsulosin, an α1A-adrenergic receptor inhibitor, is prescribed to treat benign prostatic hyperplasia in men >60 years of age, the same demographic most susceptible to abdominal aortic aneurysm. The goal of this study was to investigate the effect of tamsulosin on abdominal aortic aneurysm pathogenesis. METHODS: Abdominal aortic aneurysms were induced in WT C57BL/6 male mice (nâ¯=â¯9-18/group), using an established topical elastase abdominal aortic aneurysm model. Osmotic pumps were implanted in mice 5 days before operation to create the model, administering either low dose (0.125 µg/day tamsulosin), high dose (0.250µg/day tamsulosin), or vehicle treatments with and without topical application of elastase. Blood pressures were measured preoperatively and on postoperative days 0, 3, 7, and 14. On postoperative day 14, aortic diameter was measured before harvest. Sample aortas were prepared for histology and cytokine analysis. RESULTS: Measurements of systolic blood pressure did not differ between groups. Mice treated with the low dose of tamsulosin and with the high dose of tamsulosin showed decreased aortic diameter compared with vehicle-treated control (93% ± 24 versus 94% ± 30 versus 132% ± 24, respectively; Pâ¯=â¯.0003, Pâ¯=â¯.0003). Cytokine analysis demonstrated downregulation of pro-inflammatory cytokines in both treatment groups compared with the control (P < .05). Histology exhibited preservation of elastin in both low- and high-dose tamsulosin-treated groups (Pâ¯=â¯.0041 and Pâ¯=â¯.0018, respectively). CONCLUSION: Tamsulosin attenuates abdominal aortic aneurysm formation with increased preservation of elastin and decreased production of pro-inflammatory cytokines. Further studies are necessary to elucidate the mechanism by which tamsulosin attenuates abdominal aortic aneurysm pathogenesis.
Assuntos
Antagonistas de Receptores Adrenérgicos alfa 1/farmacologia , Aorta Abdominal/efeitos dos fármacos , Aneurisma da Aorta Abdominal/prevenção & controle , Tansulosina/farmacologia , Antagonistas de Receptores Adrenérgicos alfa 1/uso terapêutico , Animais , Aorta Abdominal/patologia , Aneurisma da Aorta Abdominal/induzido quimicamente , Aneurisma da Aorta Abdominal/patologia , Pressão Sanguínea/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Elastina/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Elastase Pancreática/toxicidade , Tansulosina/uso terapêutico , Resultado do TratamentoRESUMO
BACKGROUND: Chronic kidney disease (CKD) adversely affects bone microarchitecture and increases fracture risk. Historically, bone biopsy has been the 'gold standard' for evaluating renal bone disease but is invasive and infrequently performed. High-resolution magnetic resonance imaging (MRI) quantifies bone microarchitecture noninvasively. In patients with CKD, it has not been compared with results derived from bone biopsy or with imaging using dual energy X-ray absorptiometry (DXA). METHODS: Fourteen patients with end-stage kidney disease (ESKD) underwent MRI at the distal tibia, bone mineral density (BMD) by dual energy X-ray absorptiometry (DXA; hip and spine) and transiliac bone biopsies with histomorphometry and microcomputed tomography (micro-CT). All patients had biomarkers of mineral metabolism. Associations were determined by Spearman's or Pearson's rank correlation coefficients. RESULTS: MRI indices of trabecular network integrity, surface to curve ratio (S/C) and erosion index (EI), correlated to histomorphometric trabecular bone volume (S/C râ¯=â¯0.85, pâ¯=â¯0.0003; EI râ¯=â¯-0.82, pâ¯=â¯0.001), separation (S/C râ¯=â¯-0.58, pâ¯=â¯0.039; EI râ¯=â¯0.79, pâ¯=â¯0.0012) and thickness (S/C, râ¯=â¯0.65, pâ¯=â¯0.017). MRI EI and trabecular thickness (TbTh) also correlated to micro-CT trabecular separation (EI râ¯=â¯0.63, pâ¯=â¯0.02; TbTh râ¯=â¯-0.60, pâ¯=â¯0.02). Significant correlations were observed between histomorphometric mineralization and turnover indices and various MRI parameters. MRI-derived trabecular parameters were also significantly related to femoral neck BMD. CONCLUSIONS: This study highlights the heterogeneity of bone microarchitecture at differing skeletal sites. MRI demonstrates significant, relevant associations to important bone biopsy and DXA indices and warrants further investigation to assess its potential to non-invasively evaluate changes in bone structure and quality over time.
Assuntos
Imageamento por Ressonância Magnética/métodos , Insuficiência Renal Crônica/diagnóstico por imagem , Tíbia/diagnóstico por imagem , Microtomografia por Raio-X/métodos , Adulto , Densidade Óssea/fisiologia , Estudos de Coortes , Estudos Transversais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Diálise Renal/tendências , Insuficiência Renal Crônica/terapiaRESUMO
BACKGROUND: Cortical bone is a significant determinant of bone strength and its deterioration contributes to bone fragility. Thin cortices and increased cortical porosity have been noted in patients with chronic kidney disease (CKD), but the "Turnover Mineralization Volume" classification of renal osteodystrophy does not emphasize cortical bone as a key parameter. We aimed to assess trabecular and cortical bone microarchitecture by histomorphometry and micro-CT in patients with CKD G5 and 5D (dialysis). METHODS: Transiliac bone biopsies were performed in 14 patients undergoing kidney transplantation (n = 12) and parathyroidectomy (n = 2). Structural parameters were analysed by histomorphometry and micro-CT including trabecular bone volume, thickness (TbTh), number (TbN) and separation and cortical thickness (CtTh) and porosity (CtPo). Indices of bone remodelling and mineralisation were obtained and relationships to bone biomarkers examined. Associations were determined by Spearman's or Pearson's rank correlation coefficients. RESULTS: By micro-CT, trabecular parameters were within normal ranges in most patients, but all patients showed very low CtTh (127 ± 44 µm) and high CtPo (60.3 ± 22.5%). CtPo was inversely related to TbN (r = -0.56; p = 0.03) by micro-CT and to TbTh (r = -0.60; p = 0.024) by histomorphometry and correlated to parathyroid hormone values (r = 0.62; p = 0.021). By histomorphometry, bone turnover was high in 50%, low in 21% and normal in 29%, while 36% showed abnormal patterns of mineralization. Significant positive associations were observed between osteoblast surface, osteoclast surface, mineralization surface and bone turnover markers. CONCLUSIONS: Deterioration of cortical -microarchitecture despite predominantly normal trabecular parameters reinforces the importance of comprehensive cortical evaluation in patients with CKD.
Assuntos
Distúrbio Mineral e Ósseo na Doença Renal Crônica/diagnóstico por imagem , Distúrbio Mineral e Ósseo na Doença Renal Crônica/patologia , Osso Cortical/diagnóstico por imagem , Osso Cortical/patologia , Microtomografia por Raio-X , Adulto , Estudos Transversais , Feminino , Humanos , MasculinoRESUMO
BACKGROUND: Sphingosine-1-phosphate regulates endothelial barrier integrity and promotes cell survival and proliferation. We hypothesized that upregulation of sphingosine-1-phosphate during ex vivo lung perfusion would attenuate acute lung injury and improve graft function. METHODS: C57BL/6 mice (n = 4-8/group) were euthanized, followed by 1 hour of warm ischemia and 1 hour of cold preservation in a model of donation after cardiac death. Subsequently, mice underwent 1 hour of ex vivo lung perfusion with 1 of 4 different perfusion solutions: Steen solution (Steen, control arm), Steen with added sphingosine-1-phosphate (Steen + sphingosine-1-phosphate), Steen plus a selective sphingosine kinase 2 inhibitor (Steen + sphingosine kinase inhibitor), or Steen plus both additives (Steen + sphingosine-1-phosphate + sphingosine kinase inhibitor). During ex vivo lung perfusion, lung compliance and pulmonary artery pressure were continuously measured. Pulmonary vascular permeability was assessed with injection of Evans Blue dye. RESULTS: The combination of 1 hour of warm ischemia, followed by 1 hour of cold ischemia created significant lung injury compared with lungs that were immediately harvested after circulatory death and put on ex vivo lung perfusion. Addition of sphingosine-1-phosphate or sphingosine kinase inhibitor alone did not significantly improve lung function during ex vivo lung perfusion compared with Steen without additives. However, group Steen + sphingosine-1-phosphate + sphingosine kinase inhibitor resulted in significantly increased compliance (110% ± 13.9% vs 57.7% ± 6.6%, P < .0001) and decreased pulmonary vascular permeability (33.1 ± 11.9 µg/g vs 75.8 ± 11.4 µg/g tissue, P = .04) compared with Steen alone. CONCLUSIONS: Targeted drug therapy with a combination of sphingosine-1-phosphate + sphingosine kinase inhibitor during ex vivo lung perfusion improves lung function in a murine donation after cardiac death model. Elevation of circulating sphingosine-1-phosphate via specific pharmacologic modalities during ex vivo lung perfusion may provide endothelial protection in marginal donor lungs leading to successful lung rehabilitation for transplantation.
Assuntos
Lesão Pulmonar Aguda/prevenção & controle , Pulmão/efeitos dos fármacos , Lisofosfolipídeos/farmacologia , Perfusão/efeitos adversos , Substâncias Protetoras/farmacologia , Esfingosina/análogos & derivados , Isquemia Quente/métodos , Animais , Morte , Modelos Animais de Doenças , Transplante de Pulmão , Camundongos , Camundongos Endogâmicos C57BL , Soluções para Preservação de Órgãos/farmacologia , Esfingosina/farmacologiaRESUMO
BACKGROUND: The number of patients with end-stage pulmonary disease awaiting lung transplantation is at an all-time high, while the supply of available organs remains stagnant. Utilizing donation after circulatory death (DCD) donors may help to address the supply-demand mismatch. The objective of this study is to determine the potential donor pool expansion with increased procurement of DCD organs from patients who die at hospitals. MATERIAL AND METHODS: The charts of all patients who died at a single, rural, quaternary-care institution between August 2014 and June 2015 were reviewed for lung transplant candidacy. Inclusion criteria were age <65 y, absence of cancer and lung pathology, and cause of death other than respiratory or sepsis. RESULTS: A total of 857 patients died within a 1-year period and were stratified by age: pediatric <15 y (n = 32, 4%), young 15-64 y (n = 328, 38%), and old >65 y (n = 497, 58%). Those without cancer totaled 778 (90.8%) and 512 (59%) did not have lung pathology. This leaves 85 patients qualifying for DCD lung donation (pediatric n = 10, young n = 75, and old n = 0). Potential donors were significantly more likely to have clear chest X-rays (24.3% versus 10.0%, P < 0.0001) and higher mean PaO2/FiO2 (342.1 versus 197.9, P < 0.0001) compared with ineligible patients. CONCLUSIONS: A significant number of DCD lungs are available every year from patients who die within hospitals. We estimate the use of suitable DCD lungs could potentially result in a significant increase in the number of lungs available for transplantation.